Labmedica International November 2016 by Globetech

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DAILY CLINICAL LAB NEWS

Test Could Transform Cancer Diagnosis new blood test has been developed that could detect cancer earlier than ever before and it will change the way cancer is diagnosed, and in turn potentially save millions of lives. The assay is based on a simple blood test and it could be used to identify cancer in undiagnosed patients. It would be used to screen

Updated Guidelines for Clostridium Difficile Diagnostics ew guidelines on the best practice methods to diagnose Clostridium difficile infection (CDI) have been released. The latest document updates the original 2009 and includes recommendations concerning the use of new diagnostic tools such as nucleic acid amplification tests (NAAT).

The main objectives of this update are to summarize the currently available evidence concerning laboratory diagnosis of CDI and to formulate and revise recommendations to optimize CDI testing. This update is essential to improve the diagnosis of CDI and to improve uniformity in CDI diagnosis for surveillance purposes among

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Image: Courtesy of Stanford University Medical Center

Point-of-Care Test Diagnoses Respiratory Allergic Disorders urrently, immunoglobulin E tests for atopic asthma and other allergic conditions are performed in central laboratories, taking several weeks before results are known. Now, a newly-introduced point-of-care test provides results in minutes, leading to improved treatment conditions.

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Biomarkers Discovered for Chronic Kidney Disease Prognosis

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hronic kidney disease (CKD) is a critical health problem and CKD patients are highly prevalent throughout the world, and the number of endstage kidney disease (ESKD) patients, who need to receive cost-prohibitive kidney replacement therapy, is increasing year by year. Currently, there is no effective

ika virus has only recently gained attention due to recent large outbreaks worldwide and an easy to use nucleic acid amplification test (NAAT) could play an important role in the early detection of the infection and its patient management. Diagnosis of Zika virus has so far relied on serological methods, which are often time consuming, or nucleic acid amplification tests (NAATs) such as real-time reverse transcription

blood test which detects the point in time when the most common form of breast cancer has become resistant to treatment, could double the average time it takes for the disease to progress, from around two and a half to around six months. The test detects mutations to a gene called estrogen receptor 1, or ESR1, which indicate that receptors for the female hormone estrogen in the cancer cells that are usually driven Cont’d on page 7

INSIDE

he most frequently used first line screening test for colorectal cancer across Europe is the fecal immunochemical test (FIT), while patients with a positive score following FIT tests are then referred for colonoscopy.

Rapid Molecular Test Detects Zika Virus

Triage Colorectal Cancer Screening Test Reduces Colonoscopy Referral

Blood Test Detects New Breast Cancer Subtype

Image: A depiction of healthy lungs without asthma

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method to predict the prognosis of chronic kidney disease (CKD) patients, but scientists have now found that measuring D-amino acids, which present only trace in human, provides prognostic information of CKD. D-amino acids, the enantiomers of L-amino acids, are increasingly recognized as Cont’d on page 8

Clinical News . . . . . . . . . 4-56 IFCC News . . . . . . . . . . . . . 57 EFLM Corner . . . . . . . . . . . 62 Product News . . . . . . . 20-50 Industry News . . . . . . . . . .66 International Calendar . . . 66 PUBLISHED IN COOPERATION WITH

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Triage Colorectal Cancer Screening Test Reduces Colonoscopy Referral cont’d from cover

However, 94.8% of people who test positive with FIT do not have colorectal cancer and this means there are a significant number of unnecessary expensive and invasive colonoscopies performed, placing a severe burden on both the patient and the healthcare system. Colorectal cancer is ranked second among all newly diagnosed cancers and responsible for approximately 215,000 deaths in Europe each year. Early diagnosis is crucial as approximately 97% of bowel cancer patients caught at stage I will have an average five year survival rate and most will be cured, while if caught at stage IV the average survival rates fall to 7%. Of the 41,500 patients diagnosed with bowel cancer in the UK each year, only a very small proportion of these are diagnosed with stage I disease, 16% of men and 14% of women. As well as the increased rate of mortality with late diagnosis, there are also significant cost implications as treating late stage disease is often more costly than treating patients with early stage disease. It has been announced that a blood test, the NuQ

Triage Colorectal Cancer Screening Test (VolitionRx Ltd, Namur, Belgium; www.volitionrx.com) is expected to receive Conformité Européenne (CE) Marking in late 2016 and will be marketed commencing in early 2017. The test has demonstrated the potential to reduce colonoscopies by 25% while maintaining almost 97% detection of colorectal cancer. There are organized colorectal cancer screening programs in 14 of the 28 EU states with a further 10 states offering some form of public or privately accessible screening. Cameron Reynolds, MBA, CEO of VolitionRx, said, “Offering European healthcare systems a simple and easy to use blood test which can be used to triage FIT positive populations for colorectal cancer is very exciting as we are coming to market with something that potentially meets a pressing need in many European countries. After much market analysis, we believe that commercializing this product, a single normalized assay, is the quickest way to achieve significant revenue for our proprietary Nucleosomics platform.”

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Vol.33 No.7. Published, under license, by Globetech Media LLC; Copyright © 2016. All rights reserved. Reproduction in any form is forbidden without express permission. Opinions expressed are solely those of the authors, and do not represent an endorsement, or lack thereof, by the Publisher of any products or services. Teknopress Yayıncılık ve Ticaret Ltd. S¸ti. adına ˙Imtiyaz Sahibi: M. Geren • Yazı is¸leri Müdürü: Ersin Köklü Müs¸ ir Dervis¸ ˙Ibrahim Sok. 5/4, Esentepe, 34394 S¸is¸ li, ˙Istanbul P. K. 1, AVPIM, 34001 ˙Istanbul • E-mail: Teknopress@yahoo.com Baskı: Promat Web Ofset Tesisi • Orhangazi Mahallesi 1673. Sokak, No: 34 • 34510 Esenyurt, B. Çekmece • ˙Istanbul Yerel süreli yayındır. Yılda sekiz kere yayınlanır, ücretsiz dag˘ıtılır.

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Test Could Transform Cancer Diagnosis cont’d from cover

people who are at-risk or are asymptomatic, meaning it could detect cancer even before symptoms begin to show. Scientists at Swansea University Medical School (UK; www.swansea.ac.uk) have optimized this test over the past four years in more than 300 individuals using esophageal cancer as an example. The studies included healthy controls, pre-cancer patients and patients with cancer. The blood is stained with antibodies, which work as markers to see if there have been changes to the surface. It takes around 30 minutes to test the blood, and they estimate each test costs around GBP 35 or USD 46. The test was able to distinguish healthy volunteers and patients with a pre-cancerous condition of the esophagus, from those with cancer itself. In healthy control individuals, only a few mutated cells are detected per million red blood cells with a mean of about five per million, but in cancer patients this can rise by over 10-fold to 50-100 mutants per million. In patients undergoing chemotherapy, who are exposed to drugs that deliberately induce DNA mutations, the levels can be several hundred per million. Interestingly, these red blood cell mutations do not play a direct role in the cancer development process. They are “collateral damage” produced in circulating blood cells as a by-product of a cancer developing internally. The benefit of the blood cell mutation is that Visit us at it can be monitored in a simple, efficient, and noninvasive way. The actual test on the blood takes a MEDICA few hours to perform in the laboratory and can be 2016 done in any standard pathology department, as it Hall 7-AB01 utilizes standard laboratory equipment. Gareth Jenkins, PhD, a professor and lead investigator, said, “The test detects changes, known as mutations, in red blood cell surface proteins. These sugary proteins act as “Velcro” to stick cell recognition proteins to the cell surface. In mutated cells, the “Velcro” is missing and so the cells are “naked” for the protein of interest. Staining cells with fluorescent antibodies for the cell recognition proteins identifies normal from mutated cells which allows a mutant cell frequency to be calculated per person.” The study was presented on September 6, 2016, at the British Science Festival held at Swansea University, UK. ®

Image: Research suggests a new blood test, which is designed to spot cancer up to 10 years before symptoms appear, could be available within five years (Photo courtesy of Pablo Paul / Alamy). V

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Blood Test Detects New Breast Cancer Subtype cont’d from cover

by the hormone have evolved to stay permanently switched on without it which means hormonal treatments that block estrogen production will no longer be effective. Scientists at the Institute of Cancer Research (London, UK; www.icr.ac.uk) and their colleagues analyzed blood samples from a total of 783 women enrolled on two major phase III clinical trials of new treatments for advanced estrogen receptor positive breast cancer, which accounts for three quarters of all cases. DNA extraction was performed using the QIAamp Circulating Nucleic Acid Kit (Qiagen, Hilden, Germany; www.qiagen.com). Total free DNA was quantified from plasma using RNase P as the reference gene. For ESR1 mutation analysis, the team used commercially available multiplex droplet digital polymerase chain reaction (ddPCR) assays for the seven most common ESR1 mutations. The ddPCR was performed on a QX200 system (Bio-Rad, Hercules, CA, USA; www.bio-rad.com). A multiplex assay was considered mutation positive if at least two ESR1 mutant droplets were observed. The results obtained on the multiplex ddPCR were further characterized using uniplex ddPCR assays. The investigators found that nearly 40% of the 162 patient blood samples available, taken going into the trial, were Visit us at found to have mutations in the estrogen receptor. These women responded better MEDICA 2016 to fulvestrant, which delayed progression of the disease for 5.7 months, compared Hall 1-E26 to 2.6 months on exemestane. For women without ESR1 mutations both treatments, fulvestrant and exemestane, had the same effectiveness. The scientists also looked at a second trial that had compared treatment with fulvestrant and a placebo to fulvestrant and palbociclib. They found 25.3 %of patient blood samples had estrogen receptor mutations going into this trial. But because palbociclib targets different molecules, the patients had the same outcomes regardless of whether or not they had the mutation in the estrogen receptor. Nicholas Turner, MD, PhD, a medical oncologist and team leader said, “Our results show that breast cancer with and without ESR1 mutations are distinct subtypes that respond differently to treatment. These subtypes can be diagnosed simply and cheaply from a blood test, and should be considered for future clinical trials of advanced breast cancer to ensure patients are receiving the best treatment for their cancer. For the first time we should able to use a potentially simple test to help us pick the best treatment for women with advanced cancer after their initial treatment has failed. We do need to confirm the results in another trial before we can implement this clinically.” The study was published on September 1, 2016, in the Journal of Clinical Oncology. ®

Image: The QX200 Droplet Digital Polymerase Chain reaction (ddPCR) system (Photo courtesy of Bio-Rad).

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Biomarkers Discovered for Chronic Kidney Disease Prognosis cont’d from cover

potential biomarkers in several diseases. Nephrologists at the Osaka University Graduate School of Medicine (Osaka, Japan; www.osaka-u. ac.jp) prospectively enrolled 118 consecutive patients with CKD stages 3, 4 and 5, who were not on dialysis, from a single nephrology department between August 2005 and January 2009. Baseline blood samples from patients after an overnight fast were collected into plastic tubes to prepare plasma. The enantiomers of amino acids were quantified using the micro twodimensional high performance liquid chromatography (2D-HPLC) platform. The 4-fluoro-7-nitro-2,1,3-ben-

zoxadiazole (NBD)-derivatives of the amino acids were separated using a reversed-phase column (Shiseido, Tokyo, Japan; www.hplc.shiseido. co.jp). The retention time of NBDamino acids were determined by authentic amino acid enantiomers, and the amounts were determined by the calibration lines. All the quantitative data were obtained by the fluorescence detection. In the second dimension, HPLC-mass spectrometry (MS/MS) was used to confirm the presence of D-amino acids in the real biological matrices. API-5000 and Triple Quad 5500 (AB Sciex, Framingham, MA, USA; www.sciex.com) were used as MS/MS instruments.

The scientists found that 16 out of 21 D-amino acids were detected in blood from CKD patients. Further analyses revealed that D-Serine and D-Asparagine were associated with the progression of CKD; the risk of progression to ESKD was elevated from two- to four-fold in those with higher levels of those D-amino acids. The authors concluded that their study will provided the novel method to the clinician which can identify CKD patients at high-risk for progression to ESKD. The information of D-

Updated Guidelines for Clostridium Difficile Diagnostics cont’d from cover

European countries. Scientists working under the auspices of The European Society of Clinical Microbiology and Infectious Diseases (ESCMID, Basel, Switzerland; www.escmid.org) identified 795 new studies, of which 693 were excluded and 102 selected for review. Of these, a further 61 were excluded, leaving 41 studies to be taken forward and used to support the new guidelines. Reasons for exclusion included incorrect or inconsistent reference testing, incorrect or inconsistent index testing, incorrect sample storage and incomplete sample testing. A total of 43 studies were also considered from the original meta-analysis, of which 28 were excluded and 15 taken forward. Various laboratory assays available from commercial suppliers were assessed for their ability to diagnose CDI accurately. A reference test, typically cell cytotoxicity neutralization assay (CNNA) or toxigenic culture (TC) was used to investigate the accuracy of several tests that have emerged since 2009. These included enzyme immunoassays (EIA) that detect glutamate dehydrogenase or toxins A and B, and the new NAATs. The strongest recommendations based on the evidence from all studies in the meta-analysis were: Samples to be tested for CDI should not be limited to cases in which a physician has specifically recommended; a rectal swab can be used for testing by (toxigenic) culture, NAAT or glutamate dehydrogenase (GDH) EIA in patients with apparent ileus (inactive bowel with no discernable bowel sounds); single, standalone tests are not reliable and should not be used: a two-step algorithm is necessary. This two-step algorithm involves a combination of fast assays with follow

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amino acids would enable to select the best treatment for individual person, as well as the discovery of novel therapy. This method is also applicable to such diseases as life style-associated diseases, such as diabetes mellitus and hypertension and cardiovascular diseases, as the prognoses of these diseases are strongly influenced by CKD progression. The study was published originally published online on May 18, 2016, in the journal Scientific Reports.

up tests: Route one – the two-stage procedure should begin with either an NAAT or GDH EIA test. Negative tests should be treated as CDI negative, while positive tests should be followed up with a toxin A/B EIA test to confirm the result. Route two – the two-stage procedure should begin with both the GDH EIA test and the toxin A/B EIA test. If both are positive, CDI is likely to be present. If both are negative, CDI is unlikely to be present, however if GDH is positive and toxin A/B is negative, then the tests may optionally be followed up with an NAAT or TC test. E.J. Kuijper a professor of experimental microbiology and senior author of the study said, “The new guidelines are intended for use among medical microbiologists, gastroenterologists, infectious disease specialists and infection control practitioners. Our aim is to not only improve diagnosis of CDI, but also to standardize the diagnostic process across Europe to allow for improved surveillance of the disease.” The study was published online on July 22, 2016, in the journal Clinical Microbiology and Infection. V

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Rapid Molecular Test Detects Zika Virus contâ&#x20AC;&#x2122;d from cover

polymerase chain reaction (RT-PCR). RT-PCR requires the use of high precision instruments for thermal cycling reactions and skilled personnel for performing the complex protocol and data interpretation. Scientists at Orion Diagnostica OY (Espoo, Finland; www.oriondiagnostica. com) and a British colleague have developed an alternative NAAT, the reverse transcription strand invasion based amplification (RT-SIBA) assay for the rapid detection of Zika virus. During RT-SIBA reactions, Zika virus ribonucleic acid (RNA) is first reverse transcribed to complementary DNA (cDNA) followed by amplification and detection of cDNA under isothermal reaction conditions. SIBA relies on a recombinase-coated single-stranded invasion oligonucleotide (IO) for the separation of a complementary target duplex. They also compared the performance of RT-SIBA with real-time PCR for the detection of Visit us at Zika virus. RT-SIBA reaction was performed using the real-time PCR device, MEDICA Agilent MX3005P (Agilent Technologies, 2016 Santa Clara, CA, USA; www.agilent.com) Hall 1-D38 and parallel reactions performed using a portable fluorescence detection device, Orion GenRead, that is suitable for pointof-care or field applications. The team found that in RT-SIBA, the time to positive results for 1,000 copies of in vitro transcript RNA template was obtained after an average 17 minutes. Conversely, positive results for 1,000 copies of in vitro transcript RNA template by RT-PCR were obtained after an average 68 minutes. RT-SIBA allows for the simultaneous reverse transcription of Zika virus RNA and amplification of cDNA at the same reaction temperature. This allows for the rapid detection of Zika virus RNA within 30 minutes and was significantly faster than the RTPCR method. Yellow fever, dengue 1, West Nile, chikungunya virus or any of the 15 different unrelated microorganisms were not detected by the RT-SIBA and RT-PCR Zika assays. This may indicate that both the RT-SIBA and RT-PCR assays are specific for the detection of Zika virus RNA. The authors concluded that the rapid detection and high analytical sensitivity displayed by RT-SIBA for the detection of Zika virus, as well as tolerance to samplederived inhibition, demonstrate that the method may be a powerful molecular diagnostic tool for the detection of Zika virus. Since the method can be run on portable and relatively low-cost devices, it may be applied in the rapid response to outbreaks. The study was published on September 1, 2016, in the journal Diagnostic Microbiology and Infectious Disease. ÂŽ

Image: A Zika virus capsid, colored per chains. New research suggests the reverse transcription strand invasion based amplification (RT-SIBA) assay will enhance rapid detection of the Zika virus (Photo courtesy of Wikimedia Commons).

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Combining Two Diagnostic Tests Drastically Reduces Cancer Miss Rates he Papanicolaou (Pap) test is recommended for women between 21 and 65 years old as a screening test for cervical cancer. Women 30 and older who are negative on co-testing may wait as long as five years for their next testing, but physicians still recommend annual examinations. In recent years, high-risk human papillomavirus (hrHPV) testing for triaging atypical squamous cells of undetermined significance and co-testing with cytology have been implemented in clinical practice. However, clinical data for primary screening with human papillomavirus (HPV) testing alone are currently lacking. Scientists at the Houston Methodist Hospital (Houston, TX, USA; www.houstonmethodist.org) and their colleagues retrospectively reviewed the correlation of cytology, histology, and hrHPV testing through the use of a cytology laboratory quality assurance database with 130,648 Papanicolaou (Pap)

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tests interpreted at the BioReference Laboratories (Houston, TX, USA; www.bioreference.com) and Houston Methodist Hospital between March 1, 2013, and June 30, 2014. Among the 47,499 patients who had undergone cytology-HPV co-testing, 1,654 underwent follow-up biopsies. The sensitivities of the hrHPV and Pap tests were 80.8% and 81.2%, respectively, for detecting any type of cervicovaginal dysplasia and 91.3% and 90.9%, respectively, for high-grade cervicovaginal lesions. For the 253 biopsy-confirmed high-grade cervicovaginal lesions (cervical intraepithelial neoplasia grade 2+, adenocarcinoma in situ, or carcinoma), the false-negative rates for hrHPV and Pap tests were 8.7% and 9.1%, respectively. When they combined the tests, the teams found only three of the 253 cases were double negatives for both the Pap and hrHPV test. Therefore the false-negative rate for cytology-hrHPV co-testing was only 1.2%.

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Dina Mody, MD, director of cytopathology at Houston Methodist Hospital and co-author of the study said, “We have known that neither test is perfect and misses a certain number of cases, but we did not realize until we analyzed the data just how impactful the combination of these tests would be. The numbers tell me that Obstetrics and Gynecology specialists need to regularly offer co-testing, and woman age 30 or older need to proactively request co-testing.” The study was published in the May 2016 issue of the journal Cancer Cytopathology.

Infrared Technology Used to Screen for Minimally Invasive Colitis nflammatory bowel diseases, which include ulcerative colitis and Crohn’s disease, involve chronic inflammation of all or part of the digestive tract and can lead to life-threatening complications such as colorectal cancer. Assessing this inflammation remains a challenge, and clinical diagnosis is now achieved by colonoscopy, which uses an endoscope or flexible tube with a light and camera attached to examine the digestive tract. This technique is not ideal for an annual checkup or monitoring disease activity regularly because it is expensive, invasive and requires sedation. Scientists at Georgia State University (Atlanta, GA, USA; www.gsu.edu) have designed a minimally invasive screening method for ulcerative colitis, a debilitating gastrointestinal tract disorder, using emerging infrared technology which could be a rapid and cost-effective method for detecting disease that eliminates the need for biopsies and intrusive testing of the human body. The technique involves testing serum, the clear liquid that can be separated from clotted blood, for the increased presence of mannose, a sugar that is a marker for colitis, using Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) spectroscopy. This technology is sensitive to vibrations in the chemical bonds of the serum sample’s molecules and requires minimal sample preparation, making it a rapid diagnostic alternative. In the study, mice in two categories were tested for colitis using ATR-FTIR spectroscopy. The first group was mice with targeted deletion of the interleukin 10 (IL-10) gene, known as interleukin 10 knockout (IL10-/-) mice, which develop colitis through T helper immune cells. Disease in IL10-/mice closely resembles the physiological, histological and biochemical features of chronic colitis in humans. In the second group of mice, colitis was induced by administering Dextran Sodium Sulphate. Colitis in these mice is similar to ulcerative colitis in humans. In both groups, feces and blood samples were collected and tested. Mice in control groups were also tested. The scientists found ATR-FTIR spectroscopy was an effective tool for detecting colitis in mice serum, showing a significant increase in the levels of mannose, an indicator of colitis. The study was published on April 20, 2016, in the Journal of Biophotonics.

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Point-of-Care Test Diagnoses Respiratory Allergic Disorders dentifying patients with uncontrolled asthing is an important tool for healthcare professionals ma mediated by immunoglobulin E rein order to make informed treatment decisions mains an area of high unmet need and it within a single appointment, thus helping to enis estimated that in Europe, more than 30 million sure patients are not lost to follow-up and ultimatepeople are affected by asthma, and approximately ly improving patient management and outcomes. half of patients remain uncontrolled. The Niji System provides a platform for fast and Current immunoglobulin E tests are performed easy blood tests that could potentially be applied in central laboratories and it can take up to several across a variety of disease areas.” weeks before results are known; however point-ofcare testing (P-O-C) in the healthcare provider’s ofImage: The Niji test specific disposable cartridge fice has the potential to reduce the time from confor measuring total immunoglobulin E (IgE) from sultation to diagnosis and to treatment decision, whole blood (Photo courtesy of Novartis). leading to improved patient outcomes. Immunoglobulin E (IgE) and the total Visit us at concentration in blood (Total IgE) is an important biomarker for the diagnosis and MEDICA management of atopic asthma and other al2016 lergic conditions. Elevated IgE can be used Hall 1-D38 as a first indicator for atopic asthma in the general population. In patients with asthma, higher IgE levels are associated with increased asthma severity and reduced lung function. A novel in-office point-of-care diagnostic tool, the Niji System and Total IgE Test (Novartis, Basel, Switzerland; www.novartis. com) has recently been introduced and the test delivers quantitative total IgE levels in about 12 minutes using approximately 30μL of finger stick blood. The test has a dynamic range of 30 to 1,500 IU/mL, allowing for quick in-office diagnosis of IgEmediated allergic disorders in conjunction with other clinical findings. The Niji System does not require any sample preparation, lengthy setup, or any calibration procedures. Clinical and nonclinical performance evaluations conducted with the Niji Total IgE Test demonstrated performance comparable to currently marketed reference laboratory tests. The test may be a useful tool for healthcare professionals as an aid in the diagnosis of IgEmediated allergic disorders in conjunction with other clinical findings in their office setting. The Niji System is for use by medical professionals and consists of a small desktop analyzer, optional accessories, and test-specific disposable cartridges. It is based on a proprietary pyro-electric technology that supports the design of sensitive, rapid immunoassay tests using unprocessed capillary blood specimens. This allows the development of tests, which quantitate proteins, antibodies, drug molecules and metabolites in capillary blood and other body fluids within 10 to 15 minutes. The system is now cleared for sale within the European Union and in all countries recognizing the CE Mark and the company intends to launch the system in Europe in Q4 2016. Vas Narasimhan, MD, Global Head of Drug Development and Chief Medical Officer for Novartis, said, “point-of-care test-

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Breath Test May Improve Lung Cancer Screening and Monitoring esearchers have found that a biomarkers signature of four volatile organic compounds (VOCs) in breath facilitates earlier diagnosis of lung cancer and simple non-invasive monitoring of treatment effectiveness. Exhaled breath contains thousands of VOCs that vary in composition and pattern depending on, for example, a person’s health status. A subset of 4 carbonyl VOCs have been discovered in the exhaled breath of lung cancer patients. Being able to identify this lung cancer “signature” through a simple breath test has emerged as one of the most promising ways to diagnose the disease. Now the test is being used to monitor for disease recurrence after surgery. Erin M. Schumer, MD, MPH, Victor van Berkel, MD, PhD, and colleagues from the University of Louisville (Louisville, KY, USA; http://louisville.edu) analyzed breath samples collected before and after surgery from 31 lung cancer patients and compared

their carbonyl VOCs levels with samples from 187 healthy patients. The researchers found a significant decrease in overall carbonyl VOC levels following surgery, and 3 of the 4 carbonyl VOCs normalized after surgery, matching levels in the control group. “The rapid normalization of almost all of the four compounds after surgery provides strong evidence that they are directly produced by the tumor environment,” said Dr. Schumer, “This study confirms that the technology is accurate.” Dr. Schumer underscore the need for early detection: “We hope that breath analysis will allow us to diagnose patients with primary or recurrent lung cancer long before they suffer from symptoms, when we have more options for treating them.” Furthermore, lung cancer patients are currently followed after surgery with chest computed tomography (CT) scans, which can be inconvenient, expensive, and expose the patient to radiation.

In the simple procedure the patient blows a single breath into a specialized balloon, so breath collection can be performed in the physician’s office. The balloon is then connected to a pump that pulls the breath over a microchip, which traps the chemicals. The microchip is sent to the lab for chemical analysis, results are provided within hours. The pump is reusable; the balloon, microchip, and lab test together cost around USD 20, supporting the increasing acceptance of breath tests as a cost-effective, easy-toperform, non-invasive, and rapid tool in diagnosis and monitoring. The study, by Schumer E et al, was published June 9, 2016, in the Society of Thoracic Surgeons’s journal The Annals of Thoracic Surgery.

Novel Device Could Improve Cancer Detection irculating tumor cells (CTCs) are malignant cells that have escaped from a primary tumor into the bloodstream with the potential to form metastases at anatomically distant sites. A newly developed method to isolate cancer cells that have escaped from a tumor could soon pave the way for improved diagnosis and treatment. The simple process involves a special device that squeezes cells in a blood sample through tiny funnels, which drive the cancer cells and blood cells into separate streams based on differences in their size and softness. Scientists at University of British Columbia (Vancouver, BC, Canada; www.ubc.ca) used polydimethylsiloxane (PDMS) to fabricate the microfluidic device. Cell separation processes involved initially filling the microfluidic device using phosphate buffered saline (PBS) with 0.2% Pluronic F127 to prevent nonspecific adsorption of cells to the wall of the device. Whole blood samples from patients with metastatic castrate-resistant prostate cancer (mCRPC) and four healthy control donors were infused through the device through the sample inlet, while buffer solutions were infused through the buffer and oscillation inlets. For the assessment of the role of cell deformability in separation performance the scientists measured the cell size, suspending the cell samples in PBS and visualized using an Ti-U inverted microscope (Nikon, Tokyo, Japan; www.nikon. com) and Nikon’s CCD camera DS-2MBW. The size of CTCs and leukocytes from mCRPC patients were measured from the bright field image of the separated cells in suspension by drawing a circle on the outer edge of each target cells. At least 25 CTCs and leukocytes were measured for each mCRPC data set. The scientists were able to show in patients with metastatic castration-resistant prostate cancer, where CTCs are not significantly larger than leukocytes; CTCs can be captured based on deformability at 25× greater yield than with the conventional CellSearch system (Raritan, NJ, USA; www. cellsearchctc.com). Finally, the CTCs separated using this approach are collected in suspension and are available for downstream molecular characterization. The study was published on April 7, 2016, in the journal Small.

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Index Identifies Women at Risk for Rapid Bone Loss steoporosis is often referred to as a “silent” disease because individuals who have it experience few noticeable symptoms and this progressive condition occurs when bones grow structurally weak and become more likely to fracture or break. Biochemical markers of bone resorption and formation known as bone turnover markers (BTMs) have been available for clinical purposes for many years and in postmenopausal women, BTMs are correlated with histomorphometric indices of bone turnover, especially at sites of high remodeling. Elevated BTM levels are also associated with faster rates of bone loss and increased fracture risk. Scientists at the University of California, Los Angeles (CA, USA; www.ucla.edu) and their colleagues conducted a study to determine whether resorption and formation markers can be combined to gauge net bone formation across the

skeleton. The 685 women, who participated in the Study of Women’s Health across the Nation, were between the ages of 42 and 52. The women were either premenopausal or in early perimenopause when they enrolled in the study, and all of the participants included in this analysis had their final menstrual period during the follow-up portion of the study. The bone formation marker, serum osteocalcin (OC), was measured using the ELSA-OSTEO immunoradiometric assay as ng/mL (Cis-Bio International, Codolet, France; www.cisbio.com). The bone resorption marker, urinary N-telopeptide (UNTX), was measured using the Osteomark competitive inhibition enzyme immunoassay as nanomoles bone collagen equivalent [BCE] (Ostex International Inc, Seattle, WA, USA; www.ostex.com). Urinary creatinine was measured using the Cobas Mira autoanalyzer (Roche Diagnostics, Basel,

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Switzerland; www.roche.com). Lumbar spine (LS) and femoral neck (FN) bone mineral densities BMDs were assessed by dual-energy x-ray absorptiometry. The study was published on June 23, 2016, in the Journal of Clinical Endocrinology & Metabolism.

Plasma Mannose Levels May Indicate Diabetes Risk besity is associated with an increased risk for a wide range of morbidities, including insulin resistance (IR), type 2 diabetes (T2D), non-alcoholic fatty liver disease (NAFLD), and cardiovascular disease (CVD). Although the prevalence of obesity continues to dramatically increase worldwide, a clear understanding of the underlying molecular mechanisms involved in the progression of associated disorders is still lacking, but has recently been investigated. An international team of scientists led by those at the KTH-Royal Institute of Technology (Stockholm, Sweden; www.kth.se) used a systems biology-based approach and generated cell-specific integrated networks for liver, fat and muscle tissues. They generated cell-specific integrated networks (INs) by merging genome-scale metabolic, transcriptional regulatory and protein-protein interaction networks. They performed genome-wide transcriptomics analysis to determine the global gene expression changes in the liver and three adipose tissues from obese subjects undergoing bariatric surgery and integrated these data into the cell-specific INs. A total of 48 samples from the four tissue types of the obese subjects were sequenced using HiSeq 2000 and HiSeq 2500 systems (Illumina, San Diego, CA, USA; www.illumina.com). Measurement of plasma levels of glucose, mannose, fructose, amino acids, and a-hydroxybutyrate (AHB) was performed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) platform, based on a ACQUITY ultraperformance liquid chromatography system (UPLC, Waters Corporation, Milford, MA, USA; www.waters.com) and a Thermo-Finnigan LTQ mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA; www.thermofisher.com) operated at nominal mass resolution, which was equipped with an electrospray ionization (ESI) source and a linear ion trap (LIT) mass analyzer. The scientists found significantly high correlations between glucose and mannose and observed a significantly positive correlation between the plasma mannose levels and body mass index (BMI) and a significantly negative correlation between the plasma mannose levels and insulin sensitivity. They found that subjects with high mannose levels have a higher risk for type 2 diabetes (T2D) and that mannose can be used as a biomarker since blood mannose levels are quite stable and not influenced by recent food intake, unlike glucose levels. Adil Mardinoglu, PhD, a senior author of the study said, “We can measure mannose in the blood of lean or obese people and identify if they have increased risk for type 2 diabetes based on their mannose levels.” The study was published on June 23, 2016, in the journal Cell Metabolism.

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Biomarkers Could Provide Cancer Patients Better Survival Estimates ancer patients are often told by their doctors approximately how long they have to live, and how well they will respond to treatments, but there is a way to improve the accuracy of doctors’ predictions. A new method has been developed that could eventually lead to a way to do just that, using data about patients’ genetic sequences to produce more reliable projections for survival time and how they might respond to possible treatments. Scientists at the University of California -Los Angeles (UCLA, CA, USA; www.ucla.edu) and their colleagues have developed a method that analyzes various gene isoforms using data from ribonucleic acid (RNA) molecules in cancer specimens. These isoforms are combinations of genetic sequences that can produce an enormous variety of RNAs and proteins from a single gene. That process, called RNA sequencing, or RNAseq, reveals the presence and quantity of RNA molecules in a biological sample. In the method developed, the scientists analyzed the ratios of slightly different genetic sequences within the isoforms, enabling them to detect important but subtle differ-

ences in the genetic sequences. In contrast, the conventional analysis aggregates all of the isoforms together, meaning that the technique misses important differences within the isoforms. The scientists studied tissues from 2,684 people with cancer whose samples were part of the National Institutes of Health’s Cancer Genome Atlas, and they spent more than two years developing the algorithm for SURVIV (for “survival analysis of mRNA isoform variation”). The team has identified some 200 isoforms that are associated with survival time for people with breast cancer; some predict longer survival times, others are linked to shorter times. Armed with that knowledge, the scientists might eventually be able to target the isoforms associated with shorter survival times in order to suppress them and fight disease. They evaluated the performance of survival predictors using a metric called C-index and found that across the six different types of cancer they analyzed, their isoform-based predictions performed consistently better than the conventional gene-based predictions.

Yi Xing, PhD, an assistant professor and senior author of the study, said, “Our finding suggests that isoform ratios provide a more robust molecular signature of cancer patients in large-scale RNA-seq datasets. In cancer, sometimes a single gene produces two isoforms, one of which promotes metastasis and one of which represses metastasis.” The study was published on June 9, 2016, in the journal Nature Communications. Image: A SURVIV analysis of breast cancer isoforms developed at UCLA. Blue lines are associated with longer survival times, and magenta lines with shorter survival times (Photo courtesy of Professor Yi Xing).

Additional Hormone Measurement Reveals High Risk of Preeclampsia n additional blood test for pregnant women accurately predicts which women with high thyroid function are at risk of developing preeclampsia. The findings may help identify high-risk pregnant women and potentially avoid unnecessary treatment that carries the risk of fetal abnormalities. Preeclampsia is a condition that occurs during the second half of pregnancy, where women have high blood pressure and pass protein in their urine. It occurs in 2% to 8% of pregnancies and in some cases leads to serious complications for both mother and child, including seizures, kidney failure, hemorrhage and preterm birth. Scientists at the Erasmus University Medical Center (Rotterdam, The Netherlands; www.erasmusmc. nl) measured the hormones of 5,153 women during early pregnancy, before the 18th week, and found that women with high levels of thyroid hormone but low levels of human chorionic gonadotropin (hCG) were between three and eleven times more likely to develop preeclampsia. One of the risk factors for preeclampsia is hyperthyroidism, which can be caused by medical conditions such as Graves’ disease or toxic thyroid nodules. However, high levels of hCG, a hormone that rises naturally during pregnancy, also leads to high thyroid function but this type of pregnancy-related hyperthyroidism does not have an increased risk of preeclampsia. The combination of high-normal free thyroxine (FT4) levels with low hCG of less than 20, 000 IU/L) was associated with an 11.1-fold increased risk of preeclampsia. The combination of low thyroid-stimulating hormone (TSH) with low hCG of less than 0.000 IU/L was associated with an increased risk of preeclampsia ranging between a 9.2fold increased risk for TSH of less than 0.1 mU/L, to an 8.7-fold increased risk for TSH <5th percentile and to an 3.8-fold increased risk for TSH of

LabMedica International November/2016

less than 0.4 mU/L. The combination of high-normal FT4 and hCG of more than 20,000 or low TSH and hCG of more than 20,000 was not associated with an increased risk of preeclampsia. The authors concluded that the additional measurement of hCG in women with high normal thyroid function tests markedly improves the identification of women at high-risk of developing preeclampsia. This is likely to be due to the fact that hCG measurements allow for distinguishing physiologically high thyroid function caused by high hCG levels from pathophysiological high thyroid function due to autonomous production and/or TSH receptor stimulation antibodies. Tim I M Korevaar, MD, the lead author of the es enc r Sci o Life our Do–––– at –Y–––––ropean –– w Eu ouse! Ne areh W

study ,said, “Most pregnant women will have high thyroid hormone levels because of a natural rise in hCG, rather than an underlying thyroid condition like Grave’s Disease or toxic nodules. Doctors do not currently screen for preeclampsia, although many do measure thyroid hormone during pregnancy. Measuring hCG as well may help doctors to more accurately interpret thyroid function tests in pregnant women. Our work will potentially reassure the vast majority of patients who do not actually have an underlying thyroid condition by helping them avoid unnecessary treatment.” The study was presented at the 18th European Congress of Endocrinology held May 28 to May 31, 2016, in Munich, Germany.

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Lateral Flow Urine Assay Detects TB in HIV+ Adults uberculosis (TB) is a leading cause of death in human immunodeficiency virus (HIV)positive people and conventional sputum tests for TB take time and are not always accurate in people with HIV. A point-of-care test that does not depend on sputum evaluation, if sufficiently accurate, could help HIV-positive people who suffer high morbidity and mortality, by earlier detection of TB that may be missed by conventional sputum testing. An international review team led by the those at the Liverpool School of Tropical Medicine (UK; www.lstmed.ac.uk) have prepared a systematic review to assess the accuracy of a point-of-care urine test for diagnosing and screening for TB in people living with HIV infection. The review looked at accuracy of the lateral flow urine lipoarabinomannan assay (LF-LAM), a commercially available test that detects lipoarabinomannan (LAM), a component of mycobacterial cell walls, which is present in some people with active TB. The test is simple to carry out, requires no special equipment and provides a result within 25 minutes.

The review team examined all data published up until February 5, 2015 and included 12 studies. Six of the studies evaluated LF-LAM for TB diagnosis, looking at people with HIV and TB symptoms, while the other six evaluated the test for TB screening looking at people with HIV regardless of the presence of TB symptoms. The lateral flow urine lipoarabinomannan assay used in the studies was the Alere Determine TB LAM Ag, (Alere Inc, Waltham, MA, USA; www.alere.com), a commercially available point-of-care test for active pulmonary and extrapulmonary TB. In HIV-positive people with TB symptoms, LFLAM shows an average sensitivity and specificity of 45% and 92%. Based on these results, in 1,000 HIVpositive people where 30% (300 people) actually have TB, LF-FAM will identify 135 people with TB and miss the diagnosis in 165 with TB. For the 700 people who do not have TB, the test will correctly identify 644 people as not having TB, but will misclassify 56 as having TB. However, the sensitivity of the test is higher in HIV-positive individuals with low CD4 cell counts who are at risk of life-threat-

ening illnesses. In patients with a CD4 less than 100 cells/μL, LF-LAM sensitivity was 56% versus 26% in patients with a CD4 count of more than 100 cells/μL. Karen R. Steingart, MD, the senior author of the study said, “LF-LAM, whether used for diagnosis or screening, has low sensitivity to diagnose TB. However, and this is key, in HIV-positive individuals with low CD4 counts who are seriously ill, LF-LAM may help with the diagnosis of TB.” The study was published on May 10, 2016, in the journal Cochrane Database of Systematic Reviews. Image: The Determine TB LAM Ag test, urine pointof-care tests screens for HIV-associated TB, providing results in minutes (Photo courtesy of Alere).

Autoantibody Panel Accurately Diagnoses Early-Stage Alzheimer’s blood test based on a panel of 50 autoimmune antibodies was shown to detect individuals with mild cognitive impairment (MCI) stage Alzheimer’s disease (AD) with an overall accuracy, sensitivity, and specificity rate of 100%. Autoantibodies are abundant and ubiquitous in human sera and have been previously demonstrated as disease-specific biomarkers capable of accurately diagnosing mild-moderate stages of AD and Parkinson’s disease. In the current study, investigators at Rowan University (Stratford, NJ, USA; www.rowan.edu) sought to determine if autoantibodies could be used as biomarkers to accurately diagnose individuals with MCI that was driven by

early stages of AD pathology. To this end, they used human protein microarrays, each containing 9,486 unique human proteins, to bind serum autoantibodies. The investigators then utilized microarrays comprising the 50 autoantibody biomarkers with the highest binding affinity to analyze serum samples from 236 subjects, including 50 mild cognitive impairment (MCI) subjects with confirmed low amyloid-beta42 (AΒ42) levels. Results revealed that a small panel of bloodborne autoantibody biomarkers could be used to distinguish subjects with AD-associated MCI from age-matched and gender-matched controls with an overall accuracy of 100%. In addition, MCI subjects were successfully differentiated from those with mild-moderate AD with similar overall accuracy, suggesting that this approach might also be useful for delineation of discrete disease stages along the MCI-to-AD continuum. Finally, the panel of AD-associated MCI biomarkers was highly specific for MCI in that they accurately distinguished AD-associated MCI subjects from those with other neurodegenerative and non-neurodegenerative diseases, including early and mildmoderate Parkinson’s disease, multiple sclerosis, and early-stage breast cancer. “It is now generally believed that Alzheimer’s-related changes begin in the brain at least a decade before the emergence of telltale symptoms,” said senior author Dr. Robert Nagele, professor of osteopathic medicine at Rowan University. The study was published in the April 12, 2016, online edition of the journal Alzheimer’s & Dementia: Diagnosis, Assessment & Disease Monitoring. Image: Researchers have developed a new blood test that is able to detect the early stages of Alzheimer’s (Photo courtesy of Rowan University).

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Novel Gene Discovered for Hereditary Colon Cancer he formation of large numbers of polyps in the colon has a high probability of developing into colon cancer, if left untreated. The large-scale appearance of polyps is often due to a hereditary cause; in this case the disease can occur in multiple family members. Colon polyps form like mushroom-shaped growths from the mucosa and are several millimeters to several centimeters in size. They are benign and generally do not cause any symptoms, however, they can turn into malignant tumors or colon cancer. Physicians refer to the development of a large number of polyps in the colon as “polyposis.” Scientists at the University of Bonn (Germany; www.uni-bonn.de) and their colleagues investigated the genetic material (DNA) of polyposis patients using blood samples. They performed exome se-

quencing of leukocyte DNA from 102 unrelated individuals with unexplained adenomatous polyposis. In each patient, all of the about 20,000 protein-coding genes known were simultaneously examined. In this process, the scientists filtered the rare, possibly relevant genetic changes out of the gigantic quantity of data. They identified two unrelated individuals with differing compound-heterozygous loss-of-function (LoF) germline mutations in the mismatch-repair gene MutS Homolog 3 (MSH3) on chromosome 5. Analysis of the diseased individuals’ tumor tissue demonstrated high microsatellite instability of diand tetranucleotides (EMAST), and immunohistochemical staining illustrated a complete loss of nuclear MSH3 in normal and tumor tissue. By investigating the MSH3 gene, a clear diagnosis can be

made prospectively in some other, previously unexplained polyposis cases. Afterwards, healthy persons at risk in the family can be tested for the mutations. The study was published on August 4, 2016, in the American Journal of Human Genetics. Image: A histopathology of the colon with multiple Adenomatous polyps (Photo courtesy of Group14).

Automated Hemostasis Tests Affected by Hemolysis, Icterus, and Lipemia aboratory diagnosis is more and more prominent in modern medicine and it is commonly accepted that approximately 70% of all medical decisions are based on the laboratory results and accurate results are therefore key for appropriate diagnosis. One of the requirements for a clinical laboratory is that common interferences related to sample integrity such as hemolysis, icterus and lipemia (HIL) be evaluated with each reagent system and because of limited resources and budgetary constraints, the clinical laboratory relies on the manufacturer to document HIL estimates. Scientists at the Royal Hallamshire Hospital (Sheffield, UK; www.sth. nhs.uk) and their French colleagues collected blood samples for testing plasma-based coagulation assays and molecular hemostasis assays. They assessed the influence of HIL on prothrombin time (PT), activated partial thromboplastin time (APTT) and fibrinogen assay using a viscosity-based detection analyzer (VBDS). Interference of hemolysis was studied in two different ways: spontaneous in vitro hemolysis judged to have occurred during sample collection transport or processing and spurious hemolysis. The level of hemolysis was semiquantitatively estimated based on the measurement of hemoglobin concentration using a XN-10 (Sysmex Corporation, Kobe, Japan; www.sysmex. com). All the coagulation assays were performed using reagents and a STACompact-Max analyzer (Stago, As-

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nières sur Seine, France; www.stago. com). One reagent, Stago’s STA-Liquid Fib, titrated human calcium thrombin, was used for the measurement of fibrinogen. The scientists found that spontaneous hemolysis that occurred during sample collection and processing had no effect on PT with either a rabbit tissue factor extract or recombinant human tissue factor reagents. In contrast, addition of mechanically hemolysis cells impacted on PT for the highest hemoglobin concentration. For APTTs there was no significant difference between results in hemolysed and nonhemolysed samples. For the other two reagents studied, APTTs were statistically significantly shorter in hemolysed samples compared with nonhemolysed samples. This bias was clinically significant only for STA-PTT Automate reagent. For all three APTT reagents, the impact of hemolysis was sufficient to impact patient management decisions, and in some samples, the effects of lipemia and icterus were not clinically significant. The authors concluded that their results confirm that PT and fibrinogen are not clinically significantly affected by HIL. The APTTs of some haemolysed samples were falsely normal with one reagent and more affected than two others. Hemolysed samples should be continuously rejected. Conversely, from a clinical standpoint, lipemia and icterus did not significantly affect APTT measured with the different reagents tested in combination with a VBDS analyzer. LabMedica International November/2016

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MALDI-TOF Mass Spectrometry Differentiates Streptococcus Species t is clinically relevant to distinguish Streptococcus pneumoniae from other less virulent, members of the viridans group streptococci (VGS) and accurate species determination within the VGS and more specifically within the mitis subgroup is traditionally difficult. Matrix-assisted laser desorption ionization–timeof-flight (MALDI-TOF) mass spectrometry shows promising results for differentiation of species within the mitis group but further exploration and validation are needed. To complicate the diagnostic challenges within the VGS, in 2004, a new species within the VGS that closely resembles S. pneumoniae was described and designated as S. pseudopneumoniae. Medical microbiologists at the VU University Medical Center (Amsterdam, The Netherlands; www.vumc.com) evaluated the ability of two MALDI-TOF mass spectrometry platforms for species differentiation within the mitis subgroup. A panel consisting of 29 clinical and eight reference isolates was tested. The reference strains used included two

S. pneumoniae, two S. pseudopneumoniae, two S. mitis and two S. oralis. As a gold standard, they combined real-time polymerase chain reaction (PCR) assays targeting the Autolysin-Encoding Gene (lytA), the recombinase A (recA), and Spn9802, which were adapted from previously described methods. MALDI-TOF mass spectrometry was performed with either the Microflex platform (Bruker Daltonics; Bremen, Germany; www.bruker.com) or the Vitek MS platform (bioMérieux; Marcy l’Etiole, France; www.biomerieux-diagnostics.com). The PCR assays targeting the lytA and recA genes are specific for S. pneumoniae and S. pseudopneumoniae, respectively. The PCR assay targeting the Spn9802 fragment detects both S. pneumoniae and S. pseudopneumoniae but no other VGS. Hence, strains that tested negative in all three PCR assays were designated S. mitis/oralis. The Vitek MS correctly identified 10/11 Streptococcus pneumoniae, 13/13 Streptococcus pseudopneumoniae, and 12/13 S. mitis/oralis. The

Microflex correctly identified 9/11 S. pneumoniae, 0/13 S. pseudopneumoniae, and 13/13 S. mitis/oralis. MALDI-TOF is a powerful tool for species determination within the mitis group and diagnostic accuracy varies depending on platform and database used. The study was published in the May 2016 issue of the journal Diagnostic Microbiology and Infectious Disease. Image: The Vitek MS laser desorption ionization time-of-flight mass spectrometry for identification of bacteria platform (Photo courtesy of bioMérieux).

Blood Test Advances Diagnosis of HELLP Syndrome laboratory blood test for the diagnosis of a rare genetic red blood cell disorder also shows promise in identifying HELLP syndrome, a life-threatening high blood pressure condition affecting 1% of all pregnant women. HELLP is an acronym for hemolysis, elevated liver enzymes and low platelets and is a severe variant of pre-eclampsia whose pathogenesis remains unclear. Recent evidence and clinical similarities suggest a link to atypical hemolytic uremic syndrome, a disease of excessive activation of the alternative complement. Scientists at Johns Hopkins University School of Medicine (Baltimore, MD, USA; www.jhmi.edu) used the modified Ham test to study serum samples from 14 women with classic or atypical HELLP syn-

drome, seven women with severe preeclampsia, 11 women with normal pregnancies, and eight healthy non-pregnant women. All pregnant women were at least 23 weeks pregnant. Serum Complement C5b-9 Membrane Attack Complex levels were determined using a commercially available enzyme-linked immunosorbent assay (ELISA) kit (Quidel, San Diego, CA, USA; www.quidel.com). The modified Ham test reflects complement activation as a percentage of complement-mediated cell-killing and is thereby a functional assay. The modified Ham test was measured in an iMark Microplate Absorbance Reader (BioRad, Hercules, CA, USA; www.bio-rad.com) at 490 nm with a reference wavelength at 595 nm. The investigators found increased complement

activation, as measured by the modified Ham test, in women with classic or atypical HELLP, compared to those with normal pregnancies or those not pregnant. They observed average cell killing of 34.3% in those with classic HELLP, and 26% in atypical HELLP compared to an average 5% in those with normal pregnancies and 3.3% in those who were not pregnant. They also found that mixing serum from women with classic or atypical HELLP together with a monoclonal antibody that blocks complement, resulted in a significant decrease in the killing of cells in the modified Ham test, from about a 34% kill rate down to a 5% kill rate, the amount seen in healthy individuals. The study was published in the May 2016 issue of the journal Experimental Hematology. LabMedica International November/2016

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Immunoassay Evaluated for Quantitative Measurement of Fecal α1-Antitrypsin he measurement of the protease α1-antitrypsin (A1A) in stool specimens can be used to detect the presence of serum proteins in the gastrointestinal tract a sign of proteinlosing enteropathy (PLE). The major causes of PLE can be divided into erosive and non-erosive gastrointestinal disorders, as well as increased central venous pressure or mesenteric lymphatic obstruction and among such disorders are enteritis, Crohn disease, ulcerative colitis, and celiac disease. Scientists at the Associated Regional and University Pathologists (ARUP) Laboratories (Salt Lake City, UT, USA; www.aruplab.com) used residual stool specimens sent to ARUP laboratories for fecal A1A testing, as well as paired sera and timed stool samples obtained from healthy volunteers. A1A in stool was measured with a polyclonal antibodybased enzyme-linked immunosorbent assay (ELISA). Assay imprecision, analytical sensitivity, linearity, accuracy, analyte stability, and reference

intervals were determined. Stool A1A was measured by using the Human α1-Antitrypsin ELISA (ImmuChrom, Heppenheim, Germany; www.immuchrom.de). This assay is a polyclonal immunometric method used for quantifying the A1A extracted from stool. Signals for ELISA plates were quantified with a SPECTRAmax PLUS plate reader (Molecular Devices LLC, Sunnyvale, CA, USA; www.moleculardevices.com). Reference intervals for stool A1A and A1A clearance were established using timed stool samples and sera collected from 124 healthy volunteers (62 males, 62 females; ages 19–61 years). The investigators found that the limit of quantification of the ELISA method used was 2.0 ng/mL, and the assay was linear to 85 ng/mL. The mean recovery of A1A added to samples was 108.2% and A1A was stable in stool for a minimum of two days, seven days, and three months at room temperature, 4 °C to 8 °C and −20 °C, respectively. The upper 95th percentile reference limits for A1A in stool

and A1A clearance were 0.48 mg/g and 49 mL/day, respectively. The authors concluded that the ImmuChrom human A1A ELISA demonstrates acceptable performance characteristics for quantifying A1A in stool extracts. Furthermore, this stool ELISA may be used in combination with the TinaQuant A1A serum assay (Roche Diagnostics, Basel Switzerland; www.roche.com) to assess A1A clearance for diagnosing and monitoring PLE. The study was published in the July 2016 issue of the Journal of Applied Laboratory Medicine. Image: The SPECTRAmax PLUS plate reader (Photo courtesy of Molecular Devices).

Novel Chemiluminescent Immunoassay Detects Toxoplasma in Human Sera oxoplasmosis is common disease in humans and animals caused by the protozoan parasite Toxoplasma gondii and while this disease in healthy people is usually asymptomatic, in congenital cases and immune deficient patients it can lead to serious pathological effects. Currently, routine diagnosis of toxoplasmosis relies mainly on the use of various serological tests to detect specific antibodies in the serum samples of infected patients. The outcome of these tests depends on the antigen types and the methods of detection, and often the early recognition of the infection or precise distinction between phases of toxoplasmosis is difficult. Scientists at the Gdansk University of Technology (Poland; www.pg.edu.pl) developed new immunochemical reagents in the form of original chemilumi-

nescence (CL) labels, which enabled them to perform the immunoglobulin G (IgG) chemiluminescent immunoassay (CLIA) tests for diagnosis of human toxoplasmosis. To assess the performance of novel immunochemical reagents in the CLIA assay, SAG2-GRA1-ROP1L chimeric protein for the detection of IgG specific antibodies was utilized. The latter was previously tested as diagnostic antigen in typical enzyme-linked immunosorbent assay (ELISA). The emission profiles were simultaneously measured over 3 to 30 second periods to collect at least 95% of the CL signal, using a Centro XS3 LB-960 microplate luminometer (Berthold Technologies, Bad Wildbad, Germany; www.berthold.com). All of the sera used in the study were received from a routine toxoplasmosis screening. A total of

92 sera were analyzed, using commercial tests (Vidas Toxo-IgG II, Toxo-Screen DA, and Vidas ToxoIgM; bioMérieux, Marcy l’Etoile. France; www. biomerieux.com) and divided into the two groups: group I (IgG positive and IgM positive or negative) included 47 sera from patients with toxoplasmosis and group II (IgG and IgM negative) consisted of 45 serum samples from seronegative individuals. The diagnostic usefulness of variously acridinium ester (AE)-labeled antibodies was tested on the same day. The scientists found that the assay with the participation of AE1 conjugate expressed greater ability to distinguish the sera from patients with T. gondii infection and healthy individuals. The study was published online on May 24, 2016, in the journal Diagnostic Microbiology and Infectious Diseases.

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HEMATOLOGY REAGENTS FOR CELLCOUNTERS LABEX is a very experienced producer of reagents for various types of blood cell counters. All products are CE-marked and marketed world-wide through exclusive distributors. For further information about products, your local supplier or distribution partnership, please contact LABEX. LABEX REAGENS AB P.O. Box 22159 • SE-250 23 Helsingborg • SWEDEN Tel. +46 42 32 40 00 • Fax. +46 42 20 27 71 • gert.johansson@labex.com

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PRODUCT NEWS MOLECULAR BIOLOGY PRODUCTS

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BOTTLE-TOP DISPENSER

MICROPIPETTE

HiMedia Laboratories

Karl Hecht

SOCOREX

Products include NA extraction kits, molecular diagnostic kits (PCR/QPCR reagents), and molecular biology grade chemicals, reagents and buffers. Products also include protein purification and crystallization kits, a range of quantitative/semi-quantitative and quantitative PCR thermal cyclers.

The Dispensette S is a new bottle-top dispense for safer and convenient dispensing. It is available in several sizes, and is ideal for dispensing aggressive reagents, including concentrated bases and acids, saline solutions, as well as many organic solvents.

The Acura line includes models covering volumes from 0.5 µL up to 350 µL. The line also features an 8-channel model ranging from 10 – 100 µL. The instruments simplify sample handling, and are ideal for applications where samples and reagents are transferred to 96-well microplates.

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Novel Multiplexed Digital Immunohistochemistry Used on Tissue Sections ntratumoral heterogeneity has emerged as a critical challenge to the implementation of targeted therapeutics and immunohistochemistry (IHC) has been used to assess spatial heterogeneity of proteins; however, it has been difficult to quantify protein abundance at high multiplex and wide dynamic range. A spatially resolved, antibodybased proteomic approach has been developed with a “barcoding-potential” to quantify up to 800 targets in a single formalin-fixed paraffin-embedded (FFPE) slide. An assay has been developed capable of quantifying protein abundance in a predefined spatial region of a tissue section by labeling antibodies with photocleavable oligos, which are recognized by fluorescent barcodes and subsequently exposing them to focused UV light. Scientists at the MD Anderson Cancer Center (Houston, TX, USA; www.mdanderson.org) working with a commercial company slide-mounted FFPE tissue section bound with a multiplexed cocktail of primary antibody-oligo conjugates, and a microfluidic flow cell is attached to the slide. Using a simple modification of a standard microscope, regions of interest are identified by light or fluorescence microscopy and are sequentially illuminated with UV light to release the oligos. Following each illumination cycle, an eluent is collected and analyzed, resulting in digital counts that correspond to the abundance of each targeted protein in se-

quentially illuminated areas. The antibodies were labeled with the collaborating commercial company’s nCounter fluorescent barcodes (NanoString Technologies, Seattle, WA, USA; www.nanostring.com). The prototype includes imaging and fluidic components to capture spatial context, and existing nCounter instruments provide the quantification. Current multi-target IHC techniques involve sequential processing steps; therefore, each target addition increases the overall handling time and workload. In contrast, NanoString’s novel technology samples all analytes simultaneously to shorten tests and simplify data analysis while preserving a higher multiplexing capacity and a wider detection range. The technology is expected to be compatible with current and upcoming nCounter Vantage products for 3D Biology analysis. Joseph Beechem, PhD, Senior Vice President of R&D at NanoString said, “NanoString’s new digital IHC technology combines the high multiplexing and digital quantification of single-molecule optical barcodes with the biological insights provided by protein localization. Over the remainder of this year, our plan includes increasing the number of targets in our assays, enhancing the imaging resolution and exploring use with other 3D Biology applications.” The study was presented at the American Association for Cancer Research (AACR) annual meeting, held April 16-20, 2016, in New Orleans, LA, USA; www.aacr.org. LabMedica International November/2016

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PRODUCT NEWS CHEMICALLY DEFINED MEDIA

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PLATE ASSESSMENT SYSTEM

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LBT Innovations

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The HiCynth, in dehydrated culture media, replaces animal and plant peptones with chemically defined peptones. The HiCynth range of products reduces the variability in the raw material sources, and is environment-friendly due to an overall lower carbon footprint.

The Automated Plate Assessment System (APAS) is designed for the automated imaging, image analysis, interpretation and reporting of growth on microbiology culture plates after incubation. The APAS enables the faster diagnosis and reporting of infectious diseases.

The new generation of chicken polyclonal antibodies is designed for the two separate P100S subunits MRP8 and MRP14. By using customized antigens and a state-of-art purification technology, these antibodies fulfill the highest quality requirements and performance standards.

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Non-Invasive Prostate Cancer Test Developed Using Exosomal RNA he current screening paradigm for prostate cancer faces challenges that lead to unnecessary biopsies and, ultimately, overtreatment. The disadvantage of Prostate-Specific Antigen (PSA) testing is lack of specificity for prostate cancer resulting in both false positive and false negative results. Furthermore, PSA does not discriminate between high- and lowgrade prostate cancers. Low-grade disease could remain indolent for a long period of time and typically does not require aggressive treatment. There is a need for pre-biopsy diagnostic tools that can accurately identify high-grade prostate cancer

that requires more immediate intervention. A team of Urologists led by those at Columbia University (New York, NY, USA; www.columbia.edu) used reverse-transcriptase polymerase chain reaction (PCR), to compare the urine exosome gene expression assay with biopsy outcomes in 499 patients with prostate-specific antigen (PSA) levels of 2 to 20 ng/mL. The derived prognostic score was then validated in 1,064 patients from 22 community practice and academic urology clinic sites in the USA. Eligible participants included indolent prostate cancer (PCA)-free men, 50 years or older, scheduled for an initial or repeat-

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ed prostate needle biopsy due to suspicious digital rectal examination (DRE) findings and/or PSA levels (limit range, 2.0 to 20.0 ng/mL). The teams used the novel urinebased prostate cancer liquid biopsy test, ExoDx Prostate(IntelliScore) (Exosome Diagnostics, Inc, Cambridge, MA, USA; www.exosomedx. com) in non-DRE, first-catch urine samples. The test then analyzes the urine for three biomarkers on exosomal ribonucleic acid (exoRNA) that are expressed in men with high-grade prostate cancer. In 255 men in the training target population (median age 62 years and median PSA level 5.0 ng/mL, and initial biopsy), the urine exosome gene expression assay plus standard of care (SOC) was associated with improved discrimination between Gleason score (GS7) or greater and GS6 and benign disease. Independent validation in 519 patients urine exosome gene expression

assay plus SOC was superior to SOC. Using a predefined cut point, 138 of 519 (27%) biopsies would have been avoided, missing only 5% of patients with dominant pattern 4 high-risk GS7 disease. James McKiernan, MD, the lead author said, “The possibility of a ‘liquid biopsy’ that does not involve an invasive procedure and may be as simple as a urine test, has the potential to change the way we approach the most common cancer in men. If the predictive accuracy of the assay can be validated in further studies, it has the potential to replace the PSA test once and for all.” The study was published on March 31, 2016, in the journal JAMA Oncology. Image: A histopathology of prostate cancer showing multiple poorly formed glands with ill-defined lumina and/or incomplete nuclear complement, Gleason score 3+4 = 7 (Photo courtesy of European Urology). LabMedica International November/2016

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SEMI-AUTOMATED INSTRUMENT

FOB/TRANSFERRIN TEST

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Serum-free media is available for CHO, BHK, Vero and MDCK, most widely used mammalian cell lines, and primary cell cultures of human keratinocytes and chick embryo fibroblasts. The SFM saves time, reduces cost, improves reproducibility and maximizes product yield.

The STart Max offers new QC and calibration menus, and a large color touch screen. Features include effective traceability, the ability to archive patient results, QCs, and reagent lots. The ergonomic design improves user comfort with the incubation and measuring positions.

The Transferrin test in feces by turbidimetry (iTRA) is now available in a combo kit with i-FOB. Detection of fecal transferrin, which is more resistant to the metabolism in the intestine than hemoglobin, provides the best complementary way in early detection of colorectal cancer.

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New Blood Test Approach Identifies or Rules-Out Heart Attack wo new studies have suggested that modified algorithms using the high-sensitivity cardiac troponin I (hs-CTrop-I) assay could help more quickly identify or exclude the diagnosis of a heart attack for patients reporting to an emergency department (ED) with suspected cardiac chest pain. Current American Heart Association (AHA) guidelines recommend serial measurements of CT at presentation and 3-6 hours after symptom onset. As a result, most patients require prolonged assessment prior to safe discharge. This diagnostic approach leads to a large number of costly, potentially avoidable hospital admissions. Strategies that could safely identify a large proportion of patients suitable for discharge after a single sample of blood is taken on ED arrival would have major benefits to health care systems. In one study, Edward Carlton, PhD, North Bristol National Health Service Trust (Bristol, England;

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www.nbt.nhs.uk), and colleagues determined the diagnostic performance of low concentrations of hsCTrop-I in patients with suspected cardiac chest pain and an electrocardiogram showing no ischemia as an indicator of acute myocardial infarction (AMI; heart attack). The researchers analyzed 5 international (Australia, New Zealand, and England) observational cohort studies with outcome assessment and 30-day follow-up. A total of 3,155 patients presenting with symptoms suggestive of cardiac ischemia were included in the analysis. Eligible patients had a nonischemic electrocardiogram determined and hs-CTrop-I measured at presentation. AMI developed in 291 individuals (9.2%). hsCTrop-I concentrations that were below the limit of detection identified 19% of patients as being potentially suitable for immediate discharge, with a high diagnostic performance in excluding AMI. “To place these results in the context of absolute numbers of presenting patients, a number-needed-

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to-diagnose approach shows that, for the 1.2 ng/L cutoff level, for every 10,630 patients assessed: 1,990 would be correctly reassured that they are not having an AMI, 10 would be falsely reassured, and 8,630 would undergo further investigation, of whom 990 would ultimately receive a diagnosis of AMI. We also demonstrate that cutoff values above the lower limit of detection may not have the required diagnostic performance for clinical implementation,” the authors wrote. In the second study, Dirk Westermann, MD, University Hospital – Medical Center Hamburg-Eppendorf (Hamburg, Germany; www.uke.de/ english/index.html) and colleagues aimed to develop an algorithm for accurate and rapid exclusion and diagnosis of AMI after 1 hour. Current European Society of Cardiology guidelines recommend the use of hs-CTrop assays on admission and after 3 hours. Recent studies suggest that AMI can be diagnosed earlier than 3 hours, when values below the 99th percentile are used as cutoff values. This study investigated the application of the CTrop-I assay for the diagnosis of AMI in 1,040 patients presenting to the emergency department with acute chest pain. Results were validated in 2 independent cohorts of 4,009 patients. The researchers found that with application of a new low hs-CTropI cutoff value of 6 ng/L, the rule-out algorithm showed a high negative predictive value of 99.8% after 1 hour for AMI, allowing for accurate and rapid exclusion of AMI. The l- and 3-hour approaches yielded results that were not statistically different. Similarly, a rule-in algorithm based on hs-CTrop-I levels provided a high positive predictive value with 83%. Application of the cutoff of 6 ng/L resulted in lower follow-up mortality (1%) compared with the routinely used 99th percentile (3.7%) for this assay. “This cutoff [6 ng/L] enables a rapid triage that excludes AMI and a faster initiation of evidencebased treatment for patients diagnosed as having AMI,” the authors wrote. The two studies, by Carlton E et al and by Neumann JT et al, were published online June 1, 2016, in the journal JAMA Cardiology. LabMedica International November/2016

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PRODUCT NEWS IGG ANTIBODY TESTS

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DNA/RNA PURIFICATION SYSTEM

HEMATOLOGY ANALYZER

Euroimmun

Roche Diagnostics

Samsung

The EUROLINE range of immunoblot profiles allows determination of IgG antibodies against foods and food additives in patient serum samples. The profiles include 108 or 216 foods and food additives, and can help to establish if a food intolerance is behind chronic symptoms.

The MagNA Pure 96 is a fully automated, ultra-fast system that purifies DNA, RNA, and viral nucleic acids from a wide range of starting materials using magnetic glass particle technology. It processes up to 96 samples in less than an hour using barcoded, prefilled trays with ready-to-use reagents.

The LABGEOHC10 features a dual detection chamber, which takes 45 seconds per sample to deliver results based on 18 parameters. It performs 80 tests/hour and features an auto sample rotor to prevent sample contamination, and protect the user from contamination by blood samples.

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Genomic Study Links Migraine-Related Genes to the Vascular System esults from 22 genome-wide association (GWA) studies conducted by dozens of researchers worldwide suggest that migraine is linked to specific gene loci that are associated with some vascular disease or are involved in the regulation of vascular tone. Migraine is a debilitating neurological disorder affecting around one in seven people worldwide, but its molecular mechanisms remain poorly understood. There is some debate about whether migraine is a disease of vascular dysfunction or a result of neuronal dysfunction with secondary vascular changes. Members of the International Headache Genetics Consortium including migraine research groups from Australia, Denmark, Estonia, Finland, Germany, Iceland, the Netherlands, Norway, Spain, Sweden, the United Kingdom, and the USA carried out a major genetic study of migraine on 59,674 af-

fected subjects and 316,078 controls from 22 GWA studies. The investigators identified 44 independent single-nucleotide polymorphisms (SNPs) significantly associated with migraine risk that mapped to 38 distinct genomic loci, including 28 loci not previously reported and a locus that may be the first to be identified on the X chromosome. In subsequent computational analyses, the identified loci showed enrichment for genes expressed in vascular and smooth muscle tissues, consistent with a predominant theory of migraine that highlights links to the vascular system. “Our consortium is devoted to uncovering the genetic causes of migraine and during the past few years we have been able to identify many risk variants. Yet, in this latest, large-scale study, tens of new genetic risk factors were discovered. Because all of these variants modify the disease risk only See us at COMPAMED, Hall 8a, Stand K09

slightly, the effect could only be seen when this large amount of samples became available,” said senior author Dr. Aarno Palotie, research director at the Institute for Molecular Medicine Finland at the University of Helsinki (Finland; www.helsinki.fi) and leader of the International Headache Genetics Consortium. “We simply cannot overstate the importance of international collaboration when studying genetics of complex, common diseases.” “These interesting findings linking migraine with vascular dysfunction were generated using novel computational approaches that utilize and combine data from various international biological databases. Such datasets are invaluable in situations like this when tissue samples from patients are not readily available, underscoring the importance of data sharing,” said contributing author Dr. Benjamin Neale, instructor in medicine at the Harvard University Medical School (Boston, MA, USA; www.med.harvard.edu). The migraine genetic study was published in the June 20, 2016, online edition of the journal Nature Genetics. Image: Neurons in the brain. Researchers have identified a DNA variant that appears to regulate levels of glutamate, a chemical known as a neurotransmitter, which transports messages between neurons in the brain (Photo courtesy of Dr. Jonathan Clarke, Wellcome Images).

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Highly-Accurate Method Predicts Postpartum Diabetes estational diabetes is defined as glucose intolerance that is first identified during pregnancy and it occurs in 3% to 13% of all pregnant women, and increases a woman’s risk of developing type 2 diabetes by 20% to 50% percent within five years after pregnancy. A simple, accurate new way to predict which women with gestational diabetes will develop type 2 diabetes after delivery has been discovered which would allow health care providers to identify women at greatest risk and help motivate women to make early lifestyle changes and follow other strategies that could prevent them from developing the disease later in life. An international team of scientists working with those at the University of Toronto (ON, Canada; www.utoronto.ca) obtained fasting blood samples from 1,035 women diagnosed with gestational diabetes and enrolled in the Kaiser Permanente’s Study of Women, Infant Feeding and Type 2 Diabetes after GDM Pregnancy, also known as the SWIFT Study. The SWIFT study screened women with oral glucose tolerance tests at two months after delivery and then annually thereafter to evaluate the impact of breastfeeding and other characteristics on the development of type 2 diabetes after a pregnancy complicated by gestational diabetes. The team conducted metabolomics with baseline fasting plasma and identified 21 metabolites that significantly differed by incident type 2 diabetes (T2D) status. Machine learning optimization resulted in a decision tree modeling that predicted T2D incidence with a discriminative power of 83.0% in the training set and 76.9% in an independent testing set, being far superior to fasting plasma glucose alone. The new method may also be able to predict individuals who may develop type 2 diabetes in the general population which would be a major advance at a time when more than 300 million people suffer from the preventable form of this disease. A next-generation blood test that’s more simple and accurate than the current options could help to identify individuals who would benefit most from more timely and effective interventions to prevent type 2 diabetes. Michael B. Wheeler, PhD, a professor in the Department of Physiology and a senior author of the study said, “After delivering a baby, many women may find it very difficult to schedule two hours for another glucose test. What if we could create a much more effective test that could be given to women while they’re still in the hospital? Once diabetes has developed, it’s very difficult to reverse.” The study was published in the June 2016 issue of the journal Diabetes.

Image: Professor Michael Wheeler with Ashley St. Pierre of the Hospital for Sick Children, conducting additional tests in women with gestational diabetes to evaluate racial and ethnic differences in prediction, and investigate high-risk groups with prediabetes to learn if metabolomics will predict type 2 diabetes in the general population (Photo courtesy of the University of Toronto).

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Parkinson’s Disease Biomarker Found in Urinary Samples biomarker helps physicians predict, diagnose or monitor disease, because the biomarker corresponds to the presence or risk of disease, and its levels may change as the disease progresses. The biomarker gene, leucine-rich repeat kinase 2 (LRRK2), has been shown to play a role in hereditary Parkinson’s, and the most common of these mutations, called G2019S causes the LRRK2 kinase to add too many phosphates to itself and other proteins, but why this leads to Parkinson’s disease is not yet clear. Scientists at the University of Alabama at Birmingham (AL, USA; www.uab.edu) found that elevated phosphorylated LRRK2 predicted the risk for onset of Parkinson’s disease for people carrying a mutation in LRRK2, which is about 2% to 3% of all Parkinson’s disease patients. They tested these findings with a preliminary, 14-person cohort of urine samples from the Columbia University Movement Disorders Center. That was followed by a larger

replication study of 72 biobanked urine samples from the Michael J. Fox Foundation LRRK2 Cohort Consortium. The participants included 18 mutation carriers with Parkinson’s disease (PD), 18 mutation carriers without PD, 18 non-carriers with PD, and 18 non-carriers without PD. Exosomes were isolated from coded frozen urine by rapid thawing in a shaking heated water bath followed by differential ultracentrifugation. Exosome proteins TSG101, total LRRK2, and Ser(P)-1292 LRRK2 were measured by Western blot and LiCor Odyssey scanner analysis (Lincoln, NE,USA; www.licor.com). An exosomal pellet and a corresponding exosome protein marker (TSG101) were detected by Western blot in all lysates from all participants in the study, and LRRK2 was measurable in all preparations. The study was published originally in March 2016 in the journal Neurology and has since been expanded on June 14, 2016, in the journal Movement Disorders.

Blood Test Enables Personalized Depression Treatment blood test has been developed that accurately and reliably predicts whether depressed patients will respond to common antidepressants, which could herald a new era of personalized treatment for people with depression. Guided by this test, patients with blood inflammation above a certain threshold could be directed towards earlier access to more assertive antidepressant strategies, such as a combination of antidepressants, before their condition worsens. Scientists at King’s College London (UK; www.kcl.ac.uk) focused on two biomarkers in the blood that measure inflammation, as previous studies have already shown that elevated levels of inflammation are associated with poor response to antidepressants. They measured the quantity of two biomarkers, Macrophage Migration Inhibitory Factor (MIF) and interleukin (IL)-1 , in two independent clinical samples of depressed patients, before or after they took a range of commonly prescribed antidepressants.

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The ribonucleic acid (RNA) quantity was assessed by evaluation of the A260/280 and A260/230 ratios using a Nanodrop spectrometer (NanoDrop Technologies, Wilmington, DE, USA; www.nanodrop.com), and RNA quality was determined using an Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA; www.agilent.com). RNA samples were then stored at -80 °C until their processing for gene expression analyses. The two biomarkers examined in the study are both thought to be important in predicting how people with depression respond to antidepressants, as they are involved in several brain mechanisms relevant to depression. These include the birth of new brain cells and connections between them, as well as the death of brain cells through a process called oxidative stress. Oxidative stress occurs when the body both overproduces and then struggles to remove molecules called free radicals. The study was published on May 11, 2016, in the journal The International Journal of Neuropsychopharmacology.

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Noninvasive Test Detects Colorectal Cancer in Unscreened Patients

Image: The Cologuard testing kit for colorectal cancer (Photo courtesy of Exact Sciences).

noninvasive colorectal cancer-screening test detected the disease in patients who had previously avoided more invasive screening measures. Colorectal cancer is the second deadliest form of cancer in the USA and this year, nearly 135,000 Americans will be diagnosed with the disease and 50,000 Americans will die of it. The US Food and Drug Administration (Silver Springs, MD, USA; www.fda.gov) approved a multi-target stool DNA test (mt-sDNA) that detects the presence of red blood cells and DNA mutations that can be associated with colon cancer. In 10,000patient, prospectively conducted clinical trial the test showed 92% sensitive for detecting colorectal cancer and 42% sensitive for precancer, with a specificity of 87%. Physicians at the USMD Physician Services (Dallas, TX, USA; www.usmdinc. com) performed a retrospective medical records review of eligible patients and focused on patients at average risk for colorectal cancer, those without symptoms, a personal or family history of colorectal cancer, or polyps, which were not previously compliant with recommended guidelines for screening. During the 12month study period, from October 2014 to September 2015, USMD providers ordered 393 mt-sDNA studies, and 347 patients completed the test, achieving 88.3% compliance. Fifty-one patients, representing 14.7% of the total, tested positive by the Cologuard mt-sDNA test (Exact Sciences Corporation; Madison, WI, USA; www. cologuardtest.com) and were referred for diagnostic colonoscopies. There were 46 patients, or 90.2% of those referred, received the follow-up colonoscopies. Three patients refused the procedure and two patients did not respond to physicians’ attempts to follow up. Among the 46 patients who had follow-up colonoscopies, four were diagnosed with colon cancer. Twentyone were diagnosed with advanced adenoma, or polyps; nine had non-advanced adenoma; and 12 tested negative. Mark Prince, MD, MBA, a director of gastroenterology with USMD Physician Services, said, “Despite the availability of various colon cancer screening options, more than 40% of Americans are not getting screened. This study highlights the opportunity to expand the screening population by offering new, patient-friendly methods. We were interested to see whether the ‘real-life’ experience with Cologuard in clinical practice would be similar to the results seen in the clinical trial. Colon cancer screening saves lives. Colonoscopy is the best form of colon cancer screening, but for patients who will not have a colonoscopy, a noninvasive screening test like Cologuard is needed.” The study was presented at the American Association for Cancer Research (AACR) Annual Meeting, held April 16–20, 2016, in New Orleans (LA, USA).

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Common Circulating Cell Clusters in Cancer Patients Characterized Anew n a study of blood samples from colorectal cancer patients, researchers have now discovered the source of these cells and that they are not malignant, potentially opening up a new path to detect and inhibit the spread of cancer. Circulating cell clusters (CCCs) commonly found in the blood of cancer patients have long been regarded as malignant cells (i.e. CTCs – circulating tumor cells) that have broken off from the primary tumor, spreading cancer to other parts of the body. This was the working assumption, but due to technical challenges of separating these clusters from normal blood cells, limited research has been performed. Now a multi-institute research team, led by Dr. Min-Han Tan, a principal research scientist at Insti-

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tute of Bioengineering and Nanotechnology (IBN; Singapore; www.ibn.a-star.edu.sg) of Singapore’s A*STAR (Agency for Science, Technology & Research), has shown that these CCCs come from the blood vessels that line the tumor rather than from the tumor itself. The team includes researchers from IBN, A*STAR’s Genome Institute of Singapore, Concord Cancer Hospital, National University of Singapore, National Cancer Centre Singapore, and Singapore General Hospital. The researchers studied these CCCs at a singlecell-scale in 80 colorectal cancer patients. They first separated out the CCCs from the blood samples using a custom-designed microdevice, developed by Prof Jackie Y. Ying’s laboratory at IBN, that enables quick and efficient capture and retrieval of the CCCs. Next they used high-throughput DNA and RNA sequencing and computational modeling to determine the identity of these cells. The results showed that in colorectal cancer, these CCCs are endothelial cells from the blood vessels lining the tumor and are non-cancerous. Unexpectedly, the researchers also discovered that more endothelial CCCs were found in patients who have not received any treatment, compared to those who have received treatment, suggesting that these CCCs could be used for early-stage cancer detection. “The goal of cancer research is to understand how cancer spreads in order to curb the disease. Our institute has been focusing on evaluating cancer in a non-invasive way through blood testing using our novel microfiltration technique. Knowing exactly where these CCCs come from will lead us towards better approaches of diagnosing and treating cancer,” said Prof. Jackie Y. Ying, executive director, IBN. “Scientific orthodoxy has maintained for decades that these cell clusters commonly observed in cancer patients were malignant tumor cells. In contrast, we found that these cell clusters are not malignant, but come from the blood vessels lining

the tumor that presumably peeled off during blood flow through the tumor. This insight requires a reconsideration of decades of data, and gives scientists new opportunities to investigate and starve the cancer through drugs that manipulate the blood vessels of tumors. This method also gives physicians a new understanding and method of monitoring tumor blood supply in cancer patients receiving treatment,” said Dr Min-Han Tan. Dr Poh Koon Koh of Concord Cancer Hospital said, “I am glad that our public-private collaboration has yielded such key insights into cancer biology. Meaningful innovation comes about when focused teams are willing to challenge and disrupt existing dogmas, and the insights here allow for Singapore to develop its key technologies in the liquid biopsy domain.” The next stage of this research is to determine if the same finding applies to other types of cancer besides colorectal cancer, and to develop new liquid biopsy technologies for cancer detection and drug treatment based on these CCCs. The study, by Cima I et al, was published June 29, 2016, in the journal Science Translational Medicine. Image: Clusters of human endothelial cells captured using IBN’s custom-designed microdevice (Photo courtesy of the Institute of Bioengineering and Nanotechnology).

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Dengue Rapid Diagnostic Tests Recycled for Serotyping engue virus infection causes major public health problems in tropical and subtropical areas, but in many endemic areas, inadequate access to laboratory facilities is a major obstacle to surveillance and study of dengue epidemiology. Dengue Rapid Diagnostic Tests (RDT), in which a drop of blood is loaded onto a paper strip in a plastic cassette, are simple to use and have good diagnostic accuracy. However, four types of dengue virus circulate in most tropical areas and their patterns of circulation are of epidemiological importance since they play a role in the severity and propagation of the disease. An international team of tropical medicine specialists led by those at Mahidol University (Salaya, Thailand; www.mahidol.ac.th) collected blood samples from 99 consenting patients admitted with symptoms meeting the international criteria for dengue infection from August to November 2013. At another hospital 362 consenting patients with undifferentiated fever who tested negative by malaria RDT were enrolled from July to October 2012, and both venous whole blood and capillary whole blood from finger pricks were collected. The samples were assayed with the SD Bioline Dengue Duo RDT (Standard Diagnostics, Kyonggi-do, Korea; www. standardia.com) which is an in vitro immunochromatographic assay for the detection of dengue virus NS1 Ag and antidengue IgM/IgG antibodies in human serum, plasma, or whole blood, from finger-prick or venous blood. This test comprises a pair of test devices, a dengue NS1 Ag test on the left side, and a dengue IgM/IgG antibody (Ab) test on the right side. Dengue RNA was purified from the sample pad of the NS1 RDT loaded with virus isolates of the four serotypes, then quantified by real time reverse transcriptase polymerase chain reaction (RT-PCR). All dengue positive neat, RDT and filter paper (FP) samples were tested by real-time RT-PCR for serotyping. All four DENV serotypes were found, with the majority of patients having DENV-3 (81%; 42/52) followed by DENV-2 (10%; 5/52), DENV-4 (4%; 2/52), and DENV1(4%; 2/52) with one sample that could not be typed. There was 100% concordance between RDT and serum RT-PCR of infecting dengue serotype. The dengue serotypes at in the rural area at Salavan were mostly DENV-1 (80%; 113/142) followed by DENV-2 (12%; 17/142) and DENV-3 (4%; 6/142). The authors concluded that their technique may also permit dengue envelope sequencing for deeper molecular epidemiology analysis from RNA purified from RDTs. This could greatly increase availability of dengue epidemiological data from previously inaccessible tropical areas by facilitating dengue confirmation tests and strain identification to aid surveillance and public health interventions. The study was published on May 9, 2016, in the journal Public Library of Science Neglected Tropical Diseases.

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Image: The SD Bioline Dengue Duo Rapid Diagnostic Test (RDT) (Photo courtesy of Standard Diagnostics).

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Breath Analysis Aims to Reduce Unnecessary Antibiotic Prescriptions number of multiresistant pathogens including Acinetobacter baumannii place a heavy burden on ventilator-associated pneumonia (VAP) patients in intensive care units (ICU), and it is critically important to differentiate between bacterial infection and colonization to avoid prescribing unnecessary antibiotics. Quantitative culture of lower respiratory tract (LRT) specimens, however, requires invasive procedures, however nowadays, volatile organic compounds (VOCs) have been studied in vitro and in vivo to identify pathogen-derived biomarkers. An efficient, fast, accurate, painless and affordable test has been developed that will assist doctors in prescribing antibiotics only when the treatment is absolutely necessary. Scientists at Zhejiang University (Hangzhou, China; www.zju.edu.cn) enrolled 20 patients with A. baumannii VAP (infection group), 20 ventilated

patients with LRT A. baumannii colonization (colonization group) and 20 ventilated patients with neurological disorders, but without pneumonia or A. baumannii colonization (control group). A clinical isolate of A. baumannii strains was used for the in vitro culture experiment. For breath sample collection and gas chromatography-mass spectrometry (GC-MS) analysis the team used a GC-MS, the QP2010 Plus (Shimadzu, Kyoto, Japan; www.shimadzu.com). The detected compounds corresponding to the peaks were identified by spectra matching with the above software and Mass Spectral Library (NIST 05). The retention time and mass spectrum similarities were used to identify the compounds of interest. Breath profiles could be visually differentiated between A. baumannii cultivation in vitro and culture medium, and among in vivo groups. In the in vitro

experiment, nine compounds of interest (2,5-dimethyl-pyrazine, 1-undecene, isopentyl 3-methylbutanoate, decanal, 1,3-naphthalenediol, longifolene, tetradecane, iminodibenzyl and 3-methyl-indene) in the headspace were found to be possible A. baumannii derivations. While there were eight target VOCs (1-undecene, nonanal, decanal, 2,6,10-trimethyl-dodecane, 5-methyl-5-propyl-nonane, longifolene, tetradecane and 2-butyl-1-octanol) exhibiting characteristics of A. baumannii VAP derivations. The authors concluded that the study mainly provides a proof of concept that the direct detection of exhaled A. baumannii-derived VOCs might be adopted for an early alert of the LRT bacterial presence in ventilated ICU patients and even different parasitic states (i.e. infection and colonization) of A. baumannii. The study was published on June 7, 2016, in the Journal of Breath Research.

Urinary ELISA Kit for Alzheimer’s Diagnosis lzheimer’s disease is the most common form of dementia, a general term for memory loss and other intellectual abilities serious enough to interfere with daily life and Alzheimer’s disease (AD) accounts for 60% to 80% of dementia cases. Early, accurate diagnosis of AD is beneficial for several reasons as beginning treatment early in the disease process may help preserve daily functioning for some time, even though the underlying Alzheimer’s process cannot be stopped or reversed. Scientists at the Capital Medical University (Beijing, China; www.csc.edu.cn) have developed noninvasive and simple method for early detection of AD, which will be extremely important for the diagnosis and prognosis of AD. They aimed to develop an enzyme-linked immunosorbent assay (ELISA) kit to detect urine Alzheimer-associated neuronal thread protein (AD7C-NTP), and to evaluate its clinical value for the diagnosis of AD. The scientists collected the first morning urine specimens of 121 AD patients and 118 age-matched

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controls, and the urine AD7C-NTP levels were detected by the ELISA kit. The team synthesized immunogenic AD7C-NTP peptide fragments by the solid-phase method and used for immunizing mice or rabbits to generate anti-AD7C-NTP antibodies. The investigators reported that mouse and rabbit anti-AD7C-NTP antibody ELISA titer was found to be 1:8,000 and 1:32,000, respectively. A single band with a relative molecular mass of 41 kDa was found in human brain specimens by Western blot assay, which was identified as AD7C-NTP antibody. The urine AD7C-NTP concentration of the AD patients was higher than that of the age-matched controls, the sensitivity was 89.3% and the specificity was 84.7%. The authors concluded that they had successfully demonstrated that their newly developed urine AD7C-NTP ELISA kit has suggested potential for diagnosing AD in a Chinese population, suggesting it may be a useful diagnostic kit for detecting early AD. The study was published in the July 2016 issue of the Journal of Clinical Laboratory Analysis.

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Placental Malaria Diagnosed in Poorly Processed Placental Tissue lacental histopathology has been considered the gold standard for diagnosis of malaria during pregnancy; however, in under-resourced areas placental tissue is often improperly fixed and processed; the resulting formalin pigment is difficult to distinguish from malaria pigment. It is often problematic to diagnose malaria in pregnant women because parasitized erythrocytes sequestered in the placenta are often absent from the peripheral circulation. Microscopy of peripheral and placental blood, polymerase chain reaction (PCR), and rapid diagnostic tests have been used but vary in sensitivity and specificity. Scientists the University of North Carolina (Chapel Hill, NC, USA; www. unc.edu) and their international colleagues performed a prospective study and enrolled 182 adult women with healthy, singleton pregnancies less than 23 weeks gestational age. After delivery, a placental biopsy was collected from an incision at a healthy pericentric area of the placenta. Blood samples were collected from study participants zero to 33 days before delivery. Real-time quantitative polymerase chain reaction (qPCR) was performed on DNA extracted from filter paper-dried blood spots using a high-throughput pooling strategy. Paraffin sections of approximately 5 Îźm were stained with Giemsa according to standard histologic protocols. The stained sections were examined by two blinded microscopists under light microscopy to assess parasitemia and malaria pigment. Immunohistochemistry (IHC) was using a monoclonal antibody to Plasmodium falciparum histidine-rich protein 2 (HRP2) was carried out on 5 mm thick, formalin-fixed, paraffin-embedded sections using an automated Bond Leica IHC stainer (Leica Microsystems, Bannockburn, IL, USA; www.leicabiosystems.com). The sensitivity and specificity of individual tests were calculated using PCR or IHC as the reference standard as well as latent class analysis (LCA). The PCR (at time closest to delivery) was used as the gold standard. With PCR as the reference standard, all of the other tests had high specificities, but low sensitivities. The most sensitive other test was IHC, but it missed 50 % of the malaria infections diagnosed by PCR. The correlation between the two blinded microscopists was poor, as there was widespread formalin pigment. Using LCA, all of the tests had high specificities. The most sensitive test was IHC (67.7%), with PCR as second best (56.1%). The authors concluded that since IHC can detect parasites when they sequester in the placenta but do not circulate, while PCR detects extremely low parasitemia, the combination of both methods might

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provide a means of detecting the greatest number of infections. Therefore PCR and/or IHC are suitable diagnostics when the presence of formalin pigment substantially compromises placental histopathology. The study was published on May 10, 2016, in the Malaria Journal. Image: A photomicrograph of placental tissue revealing the presence of the malarial parasites. The blue arrows point to examples of red blood cells containing pigment and the black arrow points to an example of a pigment-containing histiocyte (Photo courtesy of Drucilla J Roberts, MD).

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PRODUCT NEWS AUTOMATED STRIP PROCESSOR

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Exposure to Toxins in Children Linked to Kidney Disease Biomarker esearchers have assessed environmental exposure to multiple toxins in children living in a region of Mexico with a high incidence of chronic kidney disease, especially among young adults. They not only detected high levels of arsenic and chromium in urine samples, they also detected elevated levels of Kidney Injury Molecule1 (KIM-1), a biomarker currently being studied as an early sign of kidney injury. Exposure to environmental toxins – such as arsenic, cadmium, chromium, lead, and other heavy metals – early in life via contaminated water or other sources often have long-term health consequences as children grow. Investigators at Brigham and Women’s Hospital (BWH; Boston, MA, USA; www.brighamandwomens.org) and Harvard Medical School (Boston, MA, USA; http://hms. harvard.edu) have found exceedingly high levels of arsenic and chromium in urine samples collected

from 107 children living in the north-central region of Mexico – levels that were even higher than exposure limits set for adults. When the team measured traditional biomarkers of kidney function – such as those measured at a routine physical exam – they did not find elevated levels of these markers. However, more sensitive and specific biomarkers of kidney injury recently qualified by the FDA for use in preclinical studies, e.g. KIM-1, were elevated in the children. KIM-1 might serve as a sensitive biomarker to screen children for kidney damage induced by environmental toxic agents. “Until now, no one has studied these children – an especially vulnerable population – to determine their risk of exposure and possible measures of kidney dysfunction,” said senior author Vishal Vaidya, PhD, of BWH and Harvard, “KIM-1 may be an early warning sign of exposure, suggesting that something may be beginning to go wrong in the epithe-

lial cells in the kidneys of these children. Many questions remain to be answered. We don’t know if this effect might be reversible, we don’t know if there are other kidney toxic contaminants such as uranium present as well. Because we don’t have follow-up data from these children we also don’t know the long-term consequences of this exposure. But this does give us our first insights into this population at a young age.” “For the first time, we’ve been able to evaluate and assess an early warning sign of kidney injury – one that may give us the ability to act in advance before there is irreversible harm,” said first author Mariana Cardenas-Gonzalez, PhD, postdoctoral fellow in the Vaidya lab. The research team also tested water samples from the children’s tap water, identifying contaminated drinking supply as the likely source of arsenic. The source of chromium, however, remains unknown so further investigation is needed. Chromium exposure can come from contaminated air, soil, water, food, or tobacco products. The study, by Cardenas-Gonzalez M et al, was published online July 15, 2016, in the journal Environmental Research. Image: New research shows children living in Mexico who are exposed to multiple toxins evidence a higher incidence of chronic kidney disease (Photo courtesy of BWH).

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Liver Cancer May Be Diagnosed from Altered Sugar Metabolism ltered gene expression that causes cancerous cells to metabolize fructose differently from healthy cells could be a significant new biomarker for diagnosing liver cancer. Dietary fructose is primarily metabolized in the liver and it has now been demonstrated that, compared with normal hepatocytes, hepatocellular carcinoma (HCC) cells markedly reduce the rate of fructose metabolism and the level of reactive oxygen species. Scientists at the University of Texas MD Anderson Cancer Center (Houston, TX, USA; www. mdanderson.org) and their colleagues discovered

Blood Test Uncovers Undiagnosed Diabetes In Hospital Patients yperglycemia is a frequent finding that can be related to physiologic stress, illness and medications, including steroids and vasopressors and glycated hemoglobin (HbA1c) correlates with the average blood glucose level over the previous eight to 12 weeks. Screening of HbA1c levels plays an important role in the diagnosis and management of diabetes mellitus in the outpatient setting but remains underused in the evaluation of hyperglycemia with undiagnosed diabetes in the inpatient setting. Scientists from the Touro University California (Vallejo, CA, USA; www.tu.edu) and the Heritage College of Osteopathic Medicine (Athens OH, USA; www.ohio.edu) reviewed the medical records of 348 patients with hyperglycemia inpatients discovered at admission in a rural community teaching hospital from June 31, 2012, to July 1, 2014. Of those patients, 50 had no known history of diabetes and 31 of them were given an HbA1C test, which measured the average blood glucose level over the previous two to three months. Fifty patients treated for hyperglycemia had medical records with no known history of diabetes (NKHD). Of the 50 patients with NKHD, 31 (62%) had an HbA1c test. Of the 31 patients tested, 6 (19%) had HbA1c levels consistent with the diagnosis of prediabetes, and 18 (58%) had levels consistent with diabetes. Seventeen (55%) of the 31 patients had a discharge diagnosis that included diabetes. Of the 19 patients with NKHD who did not have an HbA1c test, 2 (11%) received a discharge diagnosis that included diabetes. The 50 patients with NKHD had a 50% longer mean hospital stay compared with the 298 patients with known diabetes. Jay H. Shubrook, DO, the senior author of the study, said, “The study shows us that we are missing opportunities to detect diabetes and initiate treatment for those patients to help manage that disease, which can reduce their long-term cost of care and disease burden. From the osteopathic perspective of early detection equals better outcomes, it’s easy to make a case for hospital protocols to trigger an HbA1C test when hyperglycemia is detected to distinguish between transient hyperglycemia and chronic disease.” The study was published in the June 2016 issue of The Journal of the American Osteopathic Association.

that reduced fructose metabolism in liver tumor cells is caused by aberrant alternative splicing of the Ketohexokinase (Fructokinase, KHK) gene. This resulted in expression of a variety of the gene product called KHK-A, which lost the ability to process fructose. The team showed that KHK-A’s protein kinase activity enhanced tumor cell DNA and ribonucleic acid (RNA) synthesis and newly identified KHK-A as essential for liver tumor formation. Kinases are enzymes that allow cells to transfer phosphate, crucial for energy production and protein regulation. The team found that, KHK-A acts as a protein kinase, phosphorylating and activating phosphoribosyl pyrophosphate synthetase 1 (PRPS1) to promote pentose phosphate pathway-dependent de novo nucleic acid synthesis and HCC formation. Furthermore, the V-Myc Avian Myelocytomatosis Viral Oncogene Homolog (c-Myc), Heterogeneous Nuclear Ribonucleoprotein H1/2 (hnRNPH1/2)

and KHK-A expression levels and PRPS1 Thr225 phosphorylation levels correlate with each other in HCC specimens and are associated with poor prognosis for HCC. The authors concluded that their findings reveal a pivotal mechanism underlying the distinct fructose metabolism between HCC cells and normal hepatocytes and highlight the instrumental role of KHK-A protein kinase activity in promoting de novo nucleic acid synthesis and HCC development. Zhimin Lu, MD, PhD, a professor of neuro-oncology and senior author of the study said, “It is this protein kinase activity that we believe can be targeted to treat the liver tumor. Our study revealed a pivotal mechanism underlying how liver and liver tumor cells use fructose and highlight the instrumental role of the KHK-A protein in promoting tumor development.” The study was published on April 18, 2016, in the journal Nature Cell Biology.

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Mobile Phone-Based Microscopes Diagnose Intestinal Parasites andheld, mobile phonebased microscopes can be used in developing countries after minimal training of community laboratory technicians to diagnose intestinal parasites quickly and accurately. Intestinal worms affect almost two billion people world-wide, predominantly in areas with poor sanitation and unsafe water and in children, these parasites may lead to malnutrition, stunted growth and development and can lead to chronic disability, with serious health and economic consequences. An international team of scientists led by those at the University of Toronto (ON, Canada; www.utoronto.ca) trained local laboratory technicians to operate the two handheld microscopes. In total, the technicians examined stool and urine samples from 226 individuals for the detection of parasites. The accuracy of all slides was evaluated by all microscopes: the two handheld devices, as well as a conventional, “gold standard” microscope. The two portable handheld microscopes:tested were a commercial

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Newton Nm1 microscope (Newton Microscopes, Bedford, UK; http:// newtonmicroscopes.com) and a mobile phone-based CellScope (CellScope Inc, San Francisco, CA, USA; www. cellscope.com) which is essentially a smartphone with a special custom-fitted lens attached over the camera and light source, developed by engineers, to detect intestinal parasites. The scientists reported that the two handheld microscopes were very good at ruling in infections, and the Newton portable microscope was able to detect even very low-burden infections. Isaac I. Bogoch, MD, the lead investigator said, “It was heartwarming to see how well and easily these portable, handheld field microscopes were adopted and used in a rural setting. This will help us map out the areas of greatest need. Novel diagnostic approaches for common parasitic infections could have a profound impact on care of patients, as well as on public health approaches to screening in resource-poor areas.” The study was published on June 27, 2016, in the journal PLOS Neglected Tropical Diseases.

Unsuspected Bacterial Link to Bile Duct Cancer Discovered acteria in the bile duct have been identified as a potential risk factor in the development of bile duct cancer, or Cholangiocarcinoma, a rare but aggressive form of cancer with symptoms that do not present themselves at the early stages. Cholangiocarcinoma (CCA) is associated with multiple risk factors that are geographically distinct; choledocal cysts and primary sclerosing cholangitis have been implicated in the development of CCA in Western populations, while infections by the liver fluke parasite have resulted in higher incidence of CCA in Southeast Asia. An international team of scientists working with The Agency for Science, Technology and Research (Singapore; www.a-star.edu.sg) profiled 60 primary CCA tumors and matched normals, from both 28 liver fluke (Opisthorchis viverrini) associated (Ova) and 32 non-O. viverrini associated (non-OVa) cancers, using high-throughput 16S rRNA sequencing. Agencourt AMPure XP (Beckman Coulter, Brea, CA, USA; www. beckmancoulter.com) was used to purify the amplified products and purified products were visualized using

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Agilent Bioanalyzer, prepared with Agilent High Sensitivity DNA Kit (Agilent Technologies, Waldbronn, Germany; www.agilent.com). As controls for assay specificity, 16S rRNA polymerase chain reaction (PCR) was performed with extraction controls and the absence of amplification products was confirmed using Agilent Bioanalyzer. The team discovered that bile duct tissue harbored a community of diverse bacteria species. Stenotrophomonas species, previously implicated in bile duct infections, were found to be preferentially dwelling in tumor tissue compared to normal tissue of non-fluke-infected CCA patients, highlighting their potential role in development of CCA. In comparison to non-fluke-infected CCA tissues, fluke-infected CCA tissues were found to contain enteric bacteria whose metabolic outputs, bile acids and ammonia, have been previously linked to carcinogenesis, or the formation of cancers. Taken together, the results suggest a role for bile duct tissue microbiome in development of CCA and which may accordingly be used as a target for therapy. LabMedica International November/2016

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Study Improves Understanding Of E. coli H30 Epidemiology esearchers have identified populations more vulnerable to infection with pandemic E. coli strain H30, however improved testing is needed to monitor patients due to the high rate of treatment failure. Antibiotic resistance and weakened immunity only partially explain persistent infection. E. coli H30 (Sequence Type 131) begins as a subtle, hard-to-detect infection. However, even upon diagnosis and treatment with the right antibiotic, physicians often have trouble eradicating the infection, and complications can set in. The difficulty in subduing the drug-resistant E. coli H30 may also be due to an intrinsic ability that causes persistent, harmful, even deadly infections. “No other type of E. coli is causing this much widespread damage,” said co-senior author Evgeni Sokurenko, professor, University of Washington (Seattle, WA, USA; http://uw.edu), “We need to pay as much attention to it as we do to the superbug MRSA, the treatment-resistant staph infection.” It has been assumed that H30 causes opportunistic infections in the rapidly growing elderly population and others with weakened immune systems. Broad-spectrum antibiotic use may have led to drugresistance. Additionally, possible presence of as yetunknown pathogen traits could have contributed to H30’s emergence as a public health problem. A team of researchers at five USA medical centers – University of Washington, Minneapolis Veterans Affairs Health Care System, University of Minnesota, Seattle Children’s Hospital, and Group Health Cooperative – assessed the nature of H30 infections by analyzing epidemiological and medical data to explore possible associations of H30 with patient characteristics, clinical manifestations, treatment, and how patients fared. They found that individuals at greater risk for E. coli H30 infection tended to be elderly persons who had been in a healthcare facility, including longterm care residences or hospitals, who had received antibiotics, and who had underlying conditions that weakened their ability to ward off infections. The researchers also observed that patients who were later shown to have H30 were, at their first physician visit, significantly less likely to be suspected of having an infection and less likely to receive proper antibiotic. Within a month afterward, patients whose initial strain was H30 were more likely to experience severe complication. That is, H30 was strongly associated with ineffective initial therapy and diverse, later-occurring adverse effects. “It’s possible that H30 might have been hiding from the patients’ natural defenses against infection, thereby impairing the body’s attempt to clear the pathogen during treatment,” said Prof. Sokurenko, “Distinctive properties that perhaps allow H30 to act as a defenses-evading pathogen might also be associated with the delayed complications.” “H30 might have that dangerous combination of being both highly resistant to antibiotics and highly successful as a stealth pathogen,” added Prof. Sokurenko, “What makes it worse is that can go unnoticed in a patient until it causes significant damage.”

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Better methods are needed to anticipate, detect, and diagnose H30 in vulnerable patients who don’t have the usual symptoms of UTI or bloodstream infection. In addition to improved surveillance, Prof. Sokurenko would like rapid tests to become available to determine which antibiotics may or may not work for an individual patient, to help avoid antibiotic-bacteria mismatch and delayed treatment response. “Our findings reinforce the need for a more individualized medicine approach to prescribing antibiotics for E. coli infections,” he said, “Also, even if the infection seems mild, the patient should be monitored carefully so it does not progress unchecked.” Three of the study researchers have patent applications pending for E. coli H30 tests; Prof. Sokurenko is co-founder of ID Genomics. The study, by Johnson JR et al., was published

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online March 29, 2016, in the journal Clinical Infectious Diseases. Image: A scanning electron micrograph (SEM) of an E. coli bacterium of the pandemic H30 strain (Photo courtesy of the University of Washington).

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Liquid Biopsies Offer New Hope for Tracking of Ovarian Cancer

esearchers are developing a new approach that could enable early detection, monitoring, and treatment for ovarian cancer (OC) that reoccurs in patients in remission: they found that liquid biopsies from blood tests and DNA sequencing can detect OC recurrence long before a tumor reappears. OC tumors often cannot be detected until the late stages. Most patients go into remission after initial treatment, but the tumor returns 75% of the time and returning OC tumors typically do not respond to chemotherapy. The research was performed by a Mayo Clinic (Rochester, MN, USA; www.mayoclinic.org) team led by George Vasmatzis, PhD, Department of Laboratory Medicine and Pathology. “With liquid biopsies, we don’t have to wait for tumor growth to get a DNA sample,” said Dr. Vasmatzis, “This important discovery makes it possible for us to detect recurrence of the disease earlier than other diagnostic methods. We can repeat liquid biopsies to monitor the progression of the cancer.” The study was done on 10 patients in advanced stages of OC, comparing DNA from the liquid blood biopsies to tumor tissue DNA samples. Mate-pair sequencing, an inexpensive whole-exome sequencing that can reveal genetic changes that contribute to tumor growth, was used. “In this study, the blood drawn before and after surgery and the surgical tissue was used to identify DNA fragments with abnormal [chromosomal] junctions that can only be seen in this

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patient’s tumor DNA,” explained Dr. Vasmatzis, “Next-generation matepair sequencing was used to identify specific DNA changes of the tumor to create an individualized monitoring panel for liquid biopsy. This allows us to shape treatment to the individual patient rather than using a standard treatment that may not work for everyone.” Individualized tumor-specific primer panels of aberrant chromosomal junctions were identified for each case and detected by qPCR within the cell-free DNA (cfDNA). When post-surgery DNA matched that of the tumor (circulating tumor DNA [ctDNA]), patients were later found to have had a recurrence of OC. However, when the post-surgery DNA did not match the DNA of the tumor, patients were found to be in remission. The findings suggest that a primer panel for individualized chromosomal junctions derived from tumor DNA could be effective for monitoring cancer patients for relapse and therapeutic efficacy using cfDNA. The study, by Harris FR et al, was published July 20, 2016, in the journal Scientific Reports. Image: Researchers have found that DNA sequence analysis of blood samples from an initial set of ovarian cancer patients enabled detection of cancer recurrence long before a tumor reappears in analysis of tissue biopsies, suggesting that use of liquid biopsies may overcome obstacles to early detection and monitoring posed by current tumor tissuebased tests (Photo courtesy of the Mayo Clinic). LabMedica International November/2016

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Blood Test Detects Gene Mutation Associated with Lung Cancer on-small cell lung cancer (NSCLC) is the most common type of lung cancer and NSCLC tumors may shed tumor DNA into a patient’s blood, making it possible to detect specific mutations in blood samples. Testing for tumor DNA using a blood sample is also called a liquid biopsy and approval for bloodbased genetic test that can detect epidermal growth factor receptor (EGFR) gene mutations in NSCLC patients has been granted. Such mutations are present in approximately 10% to 20% of non-small cell lung cancers. Lung cancer is the leading cause of cancer-related death among men and women in the USA, and though more common in men, the number of deaths from lung cancer in women is increasing. According to the US National Cancer Institute

(Bethesda, MD, USA; www.cancer.gov), an estimated 221,200 Americans will be diagnosed with lung cancer, and 158,040 will die from the disease this year. The US Food and Drug Administration (Silver Springs MD, USA; www.fda.gov) have approved a blood-based companion diagnostic for the cancer drug Tarceva (erlotinib). The cobas EGFR Mutation Test v2 (Roche Molecular Systems, Pleasanton, CA, USA; http:// molecular.roche.com) detected the presence of specific NSCLC mutations exon 19 deletion or exon 21 (L858R) substitution mutations, in patients’ blood samples in selecting those who may benefit from treatment with Tarceva. However, if such mutations are not detected in the blood, then a tumor biopsy should be performed to determine if the NSCLC mutations are present. Insofar as the test

provides positive results, it may benefit patients who may be too ill or are otherwise unable to provide a tumor specimen for EGFR testing. Image: The cobas epidermal growth factor receptor EGFR Mutation Test v2 (Photo courtesy of Roche Molecular Systems).

Novel Test Allows for One-Step Diagnosis of HCV Infection

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he current standard in diagnosing Hepatitis C virus (HCV) infection requires two sequential steps that make it suboptimal, costly, inconvenient, time consuming, and globally not widely available or affordable. Chronic HCV infection affects approximately 170 million individuals worldwide and is associated with risk of progression to cirrhosis and hepatocellular carcinoma. Although health professional practice guidelines advocate screening for HCV infection, recent studies indicated a significant deficit in screening and diagnosis of HCV infection. Scientists at the University of California, Irvine, School of Medicine (Orange, CA, USA; www. ucirvinehealth.org) developed a novel HCV antigens enzyme immunoassay for HCV core antigen (HCVAgs EIA) and assessed its sensitivity, specificity, and utility for one-step diagnosis of viremic HCV (VHCV) infection using 365 serum specimens, including 176 without, and 189 with V-HCV infection. They confirmed presence of HCV non-structural protein 3 (NS3), NS4b, and NS5a proteins besides HCVcAg during HCV infection, and developed a novel HCV-Ags EIA via simultaneous detection of all these 4 HCV proteins. The study demonstrated that serum sample denaturation decreases the test specificity due to release of HCV-Ags sequestered in HCV-immune complexes, and should not be used in any HCV-Ags, including all the current HCVcAg assays. On the other hand, using sample non-denaturation, the HCVAgs EIA results showed 99% specificity and 100% sensitivity compared to serum anti-HCV and HCV RNA RT-PCR results. The authors concluded that the highly specific and sensitive HCV-Ags EIA they developed has the lowest limit of detection equivalent to serum HCV RNA levels of 150-250 IU/mL. Using non-denaturation of serum samples, our HCVAgs EIA reliably differentiated V-HCV infection from resolved HCV infection, accomplishes screening and diagnosis of V-HCV infection in one step. The study was published on June 6, 2016, in the journal Hepatology (www.wiley.com).

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LabMedica International

HPV Test Approved for Use with SurePath Preservative Fluid

uman papilloma virus (HPV) infections are the most common sexually transmitted infections in the USA, and HPV genotypes 16 and 18 cause approximately 70 % of cervical cancers worldwide. According to the National Cancer Institute, there will be an estimated 12,990 new cases and 4,120 deaths from cervical cancer in the USA during 2016. Prior to the approval, some laboratories used cervical cell samples collected in SurePath Preservative Fluid to run HPV tests, in lieu of collecting an additional sample in a separate collection fluid that had been approved for use with those tests. Patients who receive false negative HPV test results may not receive appropriate follow-up care, which could lead to cervical cancer progression. The US Food and Drug Administration (FDA, Silver Springs, MD, USA; www.fda.gov) approve HPV tests to be used with specific collection fluid, which store and preserve cervical cell samples for testing in the laboratory. The FDA based its approval of the Roche cobas HPV Test (Roche Molecular Systems, Basel, Switzerland; www.roche.com) with SurePath Preservative Fluid (Becton Dickinson and Company, Franklin Lakes, NJ, USA; www.bd.com) on a clinical study of 952 eligible women 21 years and older with abnormal Papanicolaou (Pap)test results. The Roche cobas HPV Test with SurePath Preservative Fluid demon-

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strated similar clinical performance when compared to a previously approved cervical sample type. Of the samples that tested positive for HPV using the Roche cobas HPV Test with SurePath, 95.4% obtained the same result as the reference sample. Of the samples that tested negative for HPV using the Roche cobas HPV Test with SurePath, 93.2% obtained the same result as the reference sample. The Roche cobas HPV Test with SurePath Preservative Fluid is approved for use with cervical cell samples obtained for a Pap test to screen women age 30 and older for HPV in order to determine whether additional follow-up and diagnostic procedures are needed. The FDA also approved the Roche cobas HPV Test with SurePath in women age 21 and older who have already had an abnormal Pap test result (borderline cellular cytology) in order to determine whether additional follow-up and diagnostic procedures are needed. The test with SurePath is also able to detect high-risk HPV genotypes 16 and 18 in the same populations of women. The Roche cobas HPV Test with SurePath is not approved as a first-line primary HPV screening test. In addition, health care professionals should use the cobas HPV Test results together with other information, such as the patient screening history and risk factors. Image: The SurePath preservative fluid kit (Photo courtesy of Becton Dickinson and Company). LabMedica International November/2016

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New Methylation Test Determines Cervical Cancer Risk novel CE-IVD-marked diagnostic triage test for use in differentiating patients’ risk of developing cervical cancer (CC) will soon be available in Europe, Middle East, and Africa for use on either clinician- or self-collected specimens. Self-sampling can streamline processes and help avoid unnecessary further medical intervention. As a follow-up assay, the test helps riskstratify patients with transforming human papillomavirus (HPV) infections. The QIAsure Methylation Test from QIAGEN N.V. (Venlo, Netherlands, and Hilden, Germany; www.qiagen.com) is a valuable addition to QIAGEN’s portfolio of leading diagnostic tools for use in women’s health and primary screening. The test is highly complementary to HPV screening tests and follows either a positive high-risk HPV test result or a finding of abnormal cells in cytology from a Pap smear. The test was presented to the public for the first time at EUROGIN 2016 (June 15-18, Salzburg, Austria), international conference for the European Research Organization on Genital Infection & Neoplasia. QIAGEN collaborated with Self-screen BV (Amsterdam, Netherlands; www.self-screen.nl), a biotech spinoff company of VU University Medical Center, in developing this test. HPV testing is generally more sensitive but less specific than cytology for detecting CC. However, the benefit of increased disease-detection using HPV as a primary test also results in many positive HPV tests in patients without underlying cervical disease. A positive HPV test result calls for followup testing to differentiate each patient’s risk based on cellular effects of the infection. With increased interest and uptake of HPV testing as a primary test, the need for a suitable triage test following a positive HPV test is increasing. Currently available triage options such as HPV genotyping or tissue-based assays often have limited clinical value. Genotyping of HPV16/18 lacks specificity, leading to false positive results; cytology lacks sensitivity, leading to missed cervical disease. The currently favored triage method for a positive HPV result is cytology. Cytology lacks sensitivity, is subjective, and skill dependent. As primary HPV testing and vaccination prevention methods are implemented, the number of patients with abnormal cells will decrease, and consequently cytology will be increasingly harder to perform. In some countries the lack of cytology infrastructure required for necessary triage is a barrier to adopting primary HPV screening. This makes a complete molecular solution highly desirable. And so the need for an accurate molecular test following a HPV-positive test result is clearly an unmet medical need. QIAsure testing accurately stratifies CC risk by detecting epigenetic changes in two specific genes implicated in CC. Specifically, it measures DNA methylation in promoter regions of host cell FAM19A4 and mir124-2 genes, which are hypermethylated in CC. In large-scale studies, the DNA markers used in this assay have demonstrated reliable additional insights into a patient’s individual risk and contributed to decisions on surveillance and treatment. By identifying hyper-methylation of

LabMedica International November/2016

these host cell genes, the QIAsure test provides highly sensitive and specific results. “QIAsure is a highly attractive and complementary addition to our leading HPV franchise”, said Thierry Bernard, senior VP and head of QIAGEN’s Molecular Diagnostics Business Area, “It creates a compelling solution for primary screening that includes the leading HPV primary screening test and leading solution for automated sample processing and molecular analysis of cervical samples.” “The QIAsure Methylation Test is an important advance for women’s health. When a woman screens positive for HPV, or cytology shows abnormal cells, she is at risk of developing CC. The QIAsure test is the next logical step to assess this risk,” said Dr. Tadd Lazarus, Chief Medical Officer, QIAGEN, “This highly sensitive, specific molecular test

identifies cancer specific epigenetic changes in cervical cells and enables the physician to assess whether the HPV infection is progressing toward cancer.” Image: A colored scanning electron micrograph (SEM) of a dividing cervical cancer cell (Photo courtesy of Wellcome Images).

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RAPID TEST

URINE CHEMISTRY ANALYZER

77 Elektronika

ELITechGroup

Sysmex

The UriSed mini offers a throughput of up to 60 tests/hour and a total measurement cycle of less than one minute. Its cost-effective operation without liquid reagents or calibrators makes it ideal for a wide range of medical and clinical settings such as hospitals, clinics, ERs and outpatient labs.

The Rapid Polymyxin NP is a rapid, reliable and cost-effective test to detect polymyxins-resistant Enterobacteriaceae. The test is easy-to-perform, very sensitive and specific, with results obtained at least 16 hours sooner than with the reference broth microdilution method.

The UC-3500 uses color CMOS sensor image technology to distinguish abnormal colors between RBCs and hemoglobin. It is capable of processing 276 samples/hour and uses the refractometry measurement method for offering accurate results.

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Heart Transplantation Recipients Unaffected by Donor Troponin Levels any transplant centers routinely reject hearts if the donor’s blood test reveals elevated levels of troponin I, a protein found in heart muscle that enters the bloodstream when there is a heart attack or other heart muscle damage.

Donors with previous heart disease are automatically excluded. Heart transplantation is one of the greatest achievements in modern medicine and patients with advanced heart failure in whom survival is measured in weeks and months are

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offered the potential for survival of equal to or greater than 10 years with excellent quality of life. Cardiologist at the Albert Einstein College of Medicine (Bronx, NY, USA; www.einstein.yu.edu) analyzed all adult heart transplant recipients at least 18 years of age, who received their transplants between January 1, 2007, and September 30, 2014. The study period coincides with the regular reporting of donor troponin levels by the United Network of Organ Sharing (UNOS). Of the 15,247 adult heart transplants performed in the USA from January 2007 to September 2014, 10 943 (71.8%) met all the inclusion/ exclusion criteria and formed the final study cohort. There were three clinically meaningful donor troponin I groups: less than 1 ng/mL, 1 to 10 ng/mL, and more than10 ng/mL and compared mortality at 30 days, one year, three years, and five years of follow-up, primary graft failure (PGF) at 30 days, and cardiac allograft vasculopathy (CAV) of more than five years of follow-up. The team set out to determine whether there are any differences in outcomes for patients who received a heart from a donor with high troponin I levels. At 30 days, one year, three years, and five years after heart transplantation, the scientists found no significant differences in survival between recipients whose donors had high troponin I levels and those whose levels were normal. There was also no association between donor troponin I levels and risk of re-

cipient death one year after transplantation. Additionally, donor troponin I levels made no difference to recipients’ incidence of primary graft failure, the loss of pumping action that occurs within 30 days of transplantation, and cardiac allograft vasculopathy, a form of heart disease that can limit long-term survival following heart transplantation. Snehal R. Patel, MD, an assistant professor of medicine and the senior author of the study, said, “A lot of focus has been on finding ways to sign up more people as organ donors, but there is also a problem in that only an average of one in three donor hearts are placed. Our study shows that transplant centers should not exclude donor hearts based solely on elevated troponin I if the organ is otherwise suitable. At our institution it has already changed how we evaluate donors, and I think this data will lead to changes nationwide.” The study was published on June 21, 2016, in the journal Circulation: Heart Failure. Image: According to new research, a blood test that leads to donor hearts being rejected may not effectively predict whether a heart transplant will succeed or fail (Photo courtesy of the AMA). LabMedica International November/2016

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LabMedica International

Lung Pathogens Detected Faster by Molecular-Based Method he genus Mycobacterium is separated into the pathogenic species Mycobacterium tuberculosis complex (MTB), Mycobacterium leprae, and Mycobacterium ulcerans, and various nontuberculous mycobacteria (NTM). New molecular-based methods are being developed to detect mycobacterial pathogens that cause pulmonary infections or tuberculosis faster than before. Time-consuming bacteria cultures no longer need to be taken from the patient samples, meaning that a suitable therapy can be started quickly. Scientists at the University of Zurich (Switzerland; www.uzh.ch) used a large-scale study with more than 6,800 patient samples to examine molecular-based methods for the detection of mycobacterial pathogens. Because many mycobacteria only grow at a very slow pace, routine detection using bacteria cultures in highly specialized and expensive high-safety laboratories takes several weeks to complete. The subsequent susceptibility test to determine the appropriate medicine also takes one to two weeks. All samples were analyzed in parallel using culture-based procedures and a modified COBAS Taq-

Man MTB test (Roche, Basel, Switzerland; www. roche.com) for direct detection of MTB and NTM.MTB polymerase chain reaction (PCR) positive specimens were screened for mutations associated with resistance to isoniazid (INH) and rifampicin (RIF) using the AID TB resistance line probe assay module 1 (AID Diagnostika GmbH, Strassberg, Germany; www.aid-diagnostika.com). Positive auramine-rhodamine microscopy results were confirmed by Ziehl-Neelsen staining. MTB molecular-based detection and culture gave concordant results for 97.7% of the specimens. NTM PCR-based detection and culture gave concordant results for 97.0% of the specimens. Defining specimens on the basis of combined laboratory data as true positives or negatives with discrepant results resolved by clinical chart reviews, the scientists calculated sensitivity, specificity, positive predictive value (PPV) and

negative predictive value (NPV) for PCR-based MTB detection as 84.7%, 100%, 100%, and 98.7%. The corresponding values for culturebased MTB detection were 86.3%, 100%, 100%, and 98.8%. PCR-based detection of NTM had a sensitivity of 84.7% compared to 78.0% of that of culture-based NTM detection. Molecular drug susceptibility testing (DST) by line-probe assay was found to predict phenotypic DST results in MTB with excellent accuracy. Peter M. Keller, MD, a senior author of the study said, “For patients and doctors, this long waiting period is an unnecessary test of their patience. By comparison, with molecular detection methods, most patients know after one or two days whether they have an infection with tuberculosis pathogens or with nontuberculous mycobacteria.” The study was published on June 16, 2016, in the journal EbioMedicine (www.ebiomedicine.com).

Urine Protein Panels Identify Prostate Cancer and Differentiate between Tumors noninvasive diagnostic test for prostate cancer is based on protein signatures that can differentiate patients from healthy individuals and those with aggressive tumors from those with less dangerous growths. Investigators at the Ontario Institute for Cancer Research (Toronto, Canada; www.oicr.on.ca) and their colleagues at the University Health Network (Toronto, Canada; www.uhn.ca) and the Eastern Virginia Medical School (Norfolk, VA, USA; www. evms.edu) used advanced proteomics techniques to generate expression information for 624 proteins obtained in urine samples. Computational analyses reduced this number by identifying significantly differentially expressed proteins and finally characterized a set of six protein biomarkers for diagnosis and a set of seven protein biomarkers for prognosis of prostate cancer. “The amazing thing about these signatures is that their rate of accuracy is as good or better than the invasive tests that are used today, with far fewer drawbacks,” said contributing author Dr. Paul Boutros, a principal investigator at the Ontario Institute for Cancer Research. “They can replace invasive, expensive, uncomfortable tests with something much easier and simpler. This type of cheap, non-invasive testing could allow patients to be screened much more frequently, allowing for more accurate monitoring of patients’ non-aggressive cancer over time, sparing patients biopsies, imaging tests and even unnecessary surgeries.” The test was described in detail in a paper published in the June 28, 2016, online edition of the journal Nature Communications.

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HEMATOLOGY ANALYZER

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Dymind Biotechnology

Edan Instruments

The EASY Line includes easy-to-use, inexpensive, rapid assays that provide evidence of a dedicated substance in a body fluid sample. It comprises a wide range of RDTs for serology, virology, DOA and STDs, and is ideal for labs requiring an economical, fast and efficient daily workflow.

The DH56 has a throughput of up to 60 samples per hour using sampling volume of only 20ul. It offers whole blood, capillary blood, and pre-dilute counting modes with the capability to flag abnormal samples and store up to 100,000 samples.

The m16 features an advanced magnetic biosensor for maximizing and shortening the clinical lab’s turnaround time, and optimizing efficiency. It uses micro-array sensor and micro-fluidic technology to provide reports in 15 minutes with samples of whole blood, plasma and serum.

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Biochip Combines Graphene Electronics And DNA Strand Displacement for Detection of Polymorphism Mutations team of bioengineers has designed an electronic biochip with potential applications for personalized medicine that can detect DNA mutations caused by single nucleotide polymorphisms (SNPs) in real time. SNPs, which are variations of a single nucleotide base, in a gene sequence are markers for a variety of human diseases. Detection of SNPs with high specificity and sensitivity is essential for effective practical implementation of personalized medicine. However, current DNA sequencing, including SNP detection, primarily uses enzyme-based methods or fluorophore-labeled assays that are time-consuming, need laboratory-scale settings, and are expensive. Previously reported electrical charge-based SNP detectors have insufficient specificity and accuracy, limiting their effectiveness. In an effort to improve the accuracy and speci-

ficity of SNP detection, investigators at the University of California, San Diego (USA; www.ucsd.edu) combined dynamic DNA nanotechnology with high resolution electronic sensing in the form of a double stranded DNA probe embedded onto a graphene field effect transistor (FET). The detection method was based on the displacement of a weakly bound DNA double strand by one containing a specific SNP. The single mismatch was detected by measuring strand displacement-induced resistance (and hence current) change in the graphene transistor. Use of large double-helix DNA strands (up to 47 bases) improved the accuracy of SNP detection by minimizing falsepositive results. “We are at the forefront of developing a fast and inexpensive digital method to detect gene mutations at high resolution – on the scale of a single nu-

cleotide change in a nucleic acid sequence,” said Dr. Lal. The SNP biosensor probe was described in detail in the June 13, 2016, online edition of the journal Proceedings of the [U.S.] National Academy of Sciences. Image: The biosensor chip - consisting of a double stranded DNA probe embedded onto a graphene transistor - electronically detects DNA SNPs (Photo courtesy of the University of California, San Diego).

Blood Coagulation Detector Helps Monitor Stroke Risk n analyzer recently developed to measure blood coagulability has the sensitivity to detect hypercoagulability associated with stroke risk in those without atrial fibrillation. Atrial fibrillation (AF) causes an irregular and sometimes fast heart rate, and is a common risk factor for stroke. A novel dielectric blood coagulometry (DBCM) has been invented for the evaluation of the coagulability and the DBCM measures the temporal change in the whole blood dielectric permittivity, which represents the aggregation of red blood cells. The CHADS2 score or CHA2DS2-Vasc predictive score are widely utilized for the risk stratification of strokes and used to guide anticoagulation therapy in patients with AF. Scientists at the Tokyo Medical and Dental University (Tokyo, Japan; www.tmd.ac.jp) and their colleagues analyzed 133 blood samples that were drawn from subjects with or without heparin ad-

ministration. A DBCM analysis was performed to find the adequate coagulation index, and to delineate its measurement range by adding recombinant human tissue factor (TF) or heparin. Then the coagulability was assessed by DBCM and conventional coagulation assays in 84 subjects without AF, who were divided into three groups by their CHADS2 score. Another 17 patients who received warfarin were also assessed by DBCM to evaluate the effect of anticoagulants. DBCM was performed using a prototype dielectric coagulometer (Sony Corporation, Tokyo, Japan; www.sony.net). The DBCM measured the dielectric permittivity in frequencies ranging from 100 Hz to 16 MHz, with sampling intervals of one minute. The measurement was completed 60 minutes after the recalcification. The dielectric permittivity was normalized compared to its initial value, and represented normalized permittivity. The result of the

DBCM was analyzed by conducting a 5-point smoothing derivative of the dielectric permittivity at 10 MHz using the linear/quadratic Savitzky-Golay filter. The investigators found that found that patients receiving warfarin had a significantly longer end of acceleration time (EAT) than those without, confirming the anticoagulation effect. They also showed that patients with a high CHADS2 score had a significantly shorter EAT that represented hypercoagulability compared with patients with lower CHADS2 scores. The authors concluded that the DBCM, is a novel highly sensitive measurement method for whole blood coagulation, can identify small changes in the coagulation status. Patients with higher CHADS2 or CHA2DS2-Vasc scores exhibited hypercoagulability without AF. The study was published on June 8, 2016 in the journal Public Library of Sciences ONE. LabMedica International November/2016

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Lipoprotein-Associated Phospholipase A2 Tests Help Diagnose Heart Disease he measurement of low-density lipoprotein cholesterol (LDL-C) or non-high-density lipoprotein cholesterol (non-HDL-C) is currently the mainstay when assessing the risk of atherosclerosis leading to cardiovascular disease. A need exists for other biomarkers of risk because not everyone with cardiovascular disease (CVD) has elevated LDL-C or non-HDL-C, and even if these markers are elevated and treatment is aggressive, there can still be residual risk that they fail to explain. A senior scientist of laboratory medicine at the Mayo Clinic (Rochester, MN, USA; www.mayo.edu) has examined an important component of atherothrombogenesis, which is inflammation, and elevated circulating biomarkers of inflammation can be used to identify individuals at higher risk of CVD. One of those markers is Lipoprotein-Associated Phospholipase A2 (Lp-PLA2), an enzyme expressed by the macrophages inside atherosclerotic plaques that circulate bound to lipoproteins. Unlike some biomarkers of inflammation, Lp-PLA2 is not an acute phase reactant, which makes it more specific for the vascular inflammation associated with CVD. The US Food and Drug Administration (FDA, Silver Springs, MD, USA; www.fda.gov) has approved two assays for Lp-PLA2 that carry the trademark name PLAC test (diaDexus, South San Francisco, CA, USA; www.diadexus.com). One measures the concentration of the protein and the other measures the enzymatic activity of the protein. The concentration assay is an enzyme-linked immunosorbent assay (ELISA) method, while the activity assay is a spectrophotometric assay run on automated chemistry analyzers. While the two assays measure the same protein, the correlation coefficients between them, somewhat surprisingly, range from 0.3 to 0.6. Laboratorians and clinicians should also be aware of several other differences between the assays. The first is evident from their respective approved patient populations: the concentration assay is cleared for coronary heart disease and stroke while the activity assay is cleared only for coronary heart disease. Last and perhaps most important, results from the ELISA test can be dramatically inaccurate as a result of pre-analytical specimen handling conditions. Leslie J. Donato, PhD, DABCC, co-director of cardiovascular laboratory medicine and co-director of the hospital clinical laboratory and point-of-care at the Mayo Clinic and the author of the review, noted that the activity assay does not display any pre-analytical effect such as storage. The only analytical vulnerability of the activity assay that is a small but progressive decrease in activity which results over the life of each lot. The laboratory director can easily monitor for this loss in signal using laboratory quality control metrics. Given the preanalytical variability that exists with the concentration assay, her group has chosen to use the activity assay when measuring Lp-PLA2. The study was published in the July 2016 edition of the journal Clinical Laboratory News.

Image: The PLAC Test activity kit for measuring Lipoprotein-Associated Phospholipase A2 (Photo courtesy of diaDexus).

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Edan Instruments

Sphere Medical

Siemens Healthineers

The i15 performs a flexible panel of tests from a single platform with automatic sample aspiration and constant temperature measurement. It provides accurate results within one minute after sample aspiration, and is suitable for a wide range of hospital and clinical applications.

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The RAPIDLab 1200 provides a comprehensive test menu, which includes full CO-oximetry and neonatal total bilirubin, to address critical care testing needs. Its microsample capability enables testing of very small samples without compromising accuracy, making it ideal for NICU monitoring.

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Human Seminal Plasma Fructose Determination Method Validated uman semen is the secretion of the male reproductive organs, containing sperm cells and seminal plasma (SP), a complex mixture of testicular, prostatic, and accessory gland secretion that provides biochemical support for ejaculate function. Assessing the quality of human semen and its fertilizing capacity therefore requires not only qualitative and quantitative analysis of spermatozoa, but also biochemical analyses of the SP. A rapid, simple, specific, and quantitative seminal fructose photometric determination by hexokinase and phosphoglucose-isomerase using a semi-automated bichromatic analyzer has been developed. Medical biochemists at the Merkur University Hospital (Zagreb, Croatia; www.mse.mef.unizg.hr) used leftover semen samples from males undergoing routine fertility evaluation whose ages ranged from

28 to 41 years, with normal semen parameters in their study. No commercial quality control material was available, so two in-house pools of SP were used for internal quality control, yielding coefficients of variation of 0.85% and 1.06% for normal and pathological values, respectively. The aim of this study was to validate the performance of the enzymatic method for SP fructose on a Beckman Coulter (BC) AU400 analyzer (Olympus Mishima, Sunto-gun, Japan; www.olympus-global.com), as well as to challenge the performance of the method regarding quality.

The determination of SP fructose by the enzymatic method is carried out indirectly via glucose. After fructose phosphorylation with hexokinase, the resulting fructose 6-phosphate is converted into glucose 6-phosphate by phosphoglucose isomerase. Glucose 6-phosphate is further oxidized by glucose 6-phosphate dehydrogenase, yielding 6-phosphogluconate and Nicotinamide adenine dinucleotide (NADH). The amount of NADH formed is stoichiometric with the amount of D-fructose and D-glucose, so the measured rate of NADH formation reflects quantitatively the amount of glucose and is equivalent to the amount of SP fructose. The methods initially compared 43 SP samples across the linearity range of 0.37 to 24.7 mmol/L. Fructose was also measured in 20 SP samples obtained from healthy volunteers without fertility problems, to confirm the previously established reference intervals of 8.3 to 27.8 mmol/L. All measured samples were within the reference interval and the results were 8.8 to 23.6 mmol/L. The study was published in the July 2016 issue of the Journal of Applied Laboratory Medicine. Image: The AU 400 fully automated, random access clinical chemistry analyzer (Photo courtesy of Olympus).

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Direct Molecular Detection of Bloodstream Infection Evaluated lood culture is the current gold standard for detecting bacteria in blood, requires at least 24 to 48 hours and has limited sensitivity if obtained during antibiotic treatment of the patient. Severe sepsis is sepsis associated with organ dysfunction, hypoperfusion, or hypotension. Rapid diagnosis and appropriate antimicrobial therapy are of major importance to decrease morbidity and mortality in patients with blood stream infections (BSI). Sepsis, severe sepsis and septic shock are associated with high mortality, ranging from 20% to 60 % depending on severity and underlying disease. Septic shock is the persistence of hypotension and perfusion abnormalities despite adequate resuscitation therapy. Scientists led by those at VU University Medical Center (Amsterdam, The Netherlands; www. vumc.com) carried out a prospective multicenter study to clinically evaluate the application of a commercial universal molecular test directly on whole blood. In total 236 samples from 166 patients with suspected sepsis were included in the study. The molecular test results were compared to blood culture, the current gold standard for detecting BSI. Because blood cultures can give false-negative results, the team performed an additional analysis to interpret the likelihood of bloodstream infection by using an evaluation based on clinical diagnosis, other diagnostic tests and laboratory parameters. Fresh EDTA blood was divided into two aliquots of 1 mL and processed according to the SepsiTest protocol (PCR-ST, Molzym, Bremen, Germany; www.molzym.com). The SepsiTest assay selectively degrades human DNA, before isolation of the microbial DNA. SepsiTest can provide a positive or negative result within four hours and needs additional sequencing to identify the microorganism, which takes another two to three hours if sequencing is available in the laboratory. The clinical interpretation of results defined the detected organism to be contaminants in 22/43 positive blood cultures (51.2 %) and 21/47 positive PCR-ST results (44.7 %). Excluding these contaminants resulted in an overall sensitivity and specificity of the PCR-ST of 66.7% and 94.4%, respectively. Of the 36 clinically relevant samples, 11 BSI were detected with both techniques, 15 BSI were detected with PCR-ST only and 10 with blood culture only. Therefore, in this study, SepsiTest detected an additional 71 % BSI compared to blood culture alone. The majority of detected microorganisms

Image: The SepsiTest allows the reliable molecular analysis of whole blood samples for bacteremia and fungemia (Photo courtesy of Molzym).

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were staphylococci in both blood culture and PCR-ST. The authors concluded that overall, PCR-ST results may influence the administration of adequate antimicrobial therapy and diminish patient’s morbidity and mortality. Although the SepsiTest PCR directly on blood is a promising technique, the input volume of blood should be increased to lower sampling error, and a faster procedure to identify the microorganism is of importance. The study was published on June 30, 2016, in the journal BMC Infectious Diseases.

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Serum Bone Alkaline Phosphatase Activity Methods Compared erum bone alkaline phosphatase is a marker of bone formation and metabolism, however, existing methods for measuring it have their limitations and their accuracy has not been determined. Alkaline phosphatase (ALP) isozymes have been used as biomarkers for liver and bone disease in the clinic and the measurement of ALP isozymes activity in serum has a high clinical value in the differential diagnosis and analysis of some diseases. Scientists at the Hamamatsu University School of Medicine (Japan; www.hama-med.ac.jp) measured serum bone ALP activity in 127 patients with liver disease using two methods, electrophoresis and chemiluminescent enzyme immunoassay (CLEIA). Of these, 52 patients had primary biliary cirrhosis, eight were affected by cholestatic cirrhosis, 13 had cholestatic hepatic disorder, and the rest were diagnosed with other liver diseases. The team used for the analysis of ALP isozymes, electrophoresis agarose gels Quick Gel ALP with Quick ALP reagent (Helena Labs, Beaumont, TX, USA; www.helena.com). In order to separate liverand bone-type ALP completely, samples were pretreated with neuraminidase. In addition, samples were pretreated with protease to separate boneand intestinal-type. Bands were scanned by densit-

ometry at a wavelength of 570 nm. ALP isozyme activity was expressed in U/L and as a percentage of total ALP activity. A one-step immunoenzymatic assay was used in this study. An automated chemiluminescence enzyme immunoassay device, known as an Access Immunoanalyzer (Beckman Coulter, Brea, CA, USA; www. beckmancoulter.com), and a special bonetype ALP (BAP) kit (also from Beckman Coulter) were used. Basic biochemical examinations of serum samples, using a chemistry analyzer (Labospect 008; Hitachi Ltd, Tokyo, Japan; www.hitachi.com), were undertaken. The investigators found that when ALP3 isozyme values were high ((%; bone-type isozyme activity as a percentage of total ALP activity), the two methods showed good correlation. However, with a decrease in ALP3 percentage levels, the correlation coefficient (R) also decreased. Starting with ALP3 (%) of less than 23%, R values markedly decreased to less than 0.5. Five outliers displayed low ALP3 (%) activity levels. Furthermore, in regard to genders, there were significant differences in total cholesterol (TC), γ-glutamyltransferase (γ-GTP), ALP and ALP3 (%) levels.

The authors concluded that the study suggests that for whole and bone-type ALP predominating (ALP2% equal to or less than ALP3%) samples, electrophoresis and CLEIA methods show good correlation. However, the accuracy of electrophoresis needs to be evaluated with further when patient samples under certain conditions. The study was published on June 9, 2016, in the journal Clinica Chimica Acta. Image: The Access 2 immunoassay system (Photo courtesy of Beckman Coulter).

LAMP Assay Developed for Threadworms in Stool and Urine trongyloides stercoralis, commonly known as threadworm, is a nematode globally distributed but mainly endemic in tropical and subtropical regions and is the chief causative agent of human strongyloidiasis. The diagnosis of strongyloidiasis is suspected when clinical signs and symptoms, eosinophilia or serologic findings are observed, but definitive diagnosis is accomplished by parasitological examination of stool samples allowing the morphological identification of S. stercoralis, including direct smear in saline,

the spontaneous sedimentation method, and centrifugation. Scientists at the University of Salamanca (Salamanca, Spain; www.usal.es) and their colleagues developed, a molecular assay using loop mediated isothermal amplification (LAMP) methodology as a simple, sensible and robust method for the detection of S. venezuelensis DNA in a well-established rodent model infection in both stool and urine samples. The LAMP assay was also successfully evaluated in patients´ stool samples. The LAMP assay

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(Strong-LAMP) is a useful molecular tool in a strongyloidiasis infection model and could be a potential field-friendly diagnostic test in a clinical setting, following further validation. Stool samples were collected from 12 patients who showed significant levels of immunoglobulin E (IgE), eosinophilia or other symptoms suggestive of disease. Stool samples were examined after arrival by qualified laboratory technicians. Eleven of these 12 stool samples were subjected to different parasitological methods as screening tests for strongyloidiasis, including microscopic examination (MOE) for the presence of rabditiform larvae in direct fecal smears, agar plate culture (APC) or Harada-Mori´s filter paper culture method (HMM). DNA was extracted from the human stool samples and tested by an optimized real-time polymerase chain reaction. To evaluate the ability of the LAMP assay designed to amplify S. venezuelensis DNA in real samples, the team used DNA extracted from the pooled feces and urine samples taken daily from each experimentally infected group of rats with different doses. All patients´ stool samples were tested by RTPCR and LAMP to compare results. The RT-PCR resulted positive in 6/7 patients´ stool samples with confirmed strongyloidiasis by parasitological tests previously applied. In addition, a positive result was obtained in a sample to which no parasitological test could be performed; negative results were obtained in negative parasitological samples for S. stercoralis. The study was published on July 14, 2016, in the journal Public Library of Science Neglected Tropical Diseases. LabMedica International November/2016

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High-Sensitivity Micro LC-MS/MS Assay Developed for Serum Estradiol here are considerable demands to accurately measure estradiol (E2) at low concentrations of less than 20 pg/mL in postmenopausal women, men, pediatric patients, and patients receiving breast cancer treatment. Automated immunoassays are the most widely used techniques for E2measurements in clinical laboratories; however, they suffer from poor accuracy and precision at low physiological E2 concentrations. A high-sensitivity, underivatized method has been developed using micro liquid chromatography–mass spectrometry (LC-MS/MS) to reliably measure E2 concentrations below 5 pg/mL by the use of low sample volume. Scientists at the University of Chicago (IL, USA; www.uchicago.edu) used samples for method comparison from

leftover samples retrieved from the clinical chemistry laboratory. A total of 290 L of sample was mixed with internal standard (IS), E2-d4, and extracted with a mixture of hexane/ethyl acetate (90/10) (v/v). After extraction, sample was separated by an Eksigent Ekspert micro LC 200 system with a flow rate of 35 L/min in a total run time of 3.5 minutes and detected by a QTRAP 6500 mass spectrometer (Sciex, Framingham, MA, USA; www. sciex.com) in a negative mode using transitions: 271/145 (quantifier) and 271/143 (qualifier). In this method, it was crucial to use HPLC columns with stability at a pH >10. E2 measurements by LC-MS/MS were compared with the Cobas 8000 E2 II automated immunoassay (Roche Diagnostics, Basel, Switzerland; www. roche.com ) using 42 plasma samples

collected in lithium heparin tubes and with a laboratory-developed indirect radioimmunoassay using 38 serum samples collected in plain serum tubes. The LCMS/MS method showed good correlation with E2 RIA and modest correlation with E2 Roche Cobas automated immunoassay. The validation study demonstrated broad linear ranges 3.0 to 820 pg/mL. Total precision was below 15% at all quality control (QC) levels, and limit of quantification (LOQ) was 3.0 pg/mL. The study was published in the July 2016 issue of the Journal of Applied Laboratory Medi-

cine. Image: The Eksigent Ekspert micro liquid chromatography (LC) 200 system (Photo courtesy of Sciex).

Brain Infections Diagnosed With Next-Generation Sequencing he feasibility of next-generation sequencing (NGS) microbiome approaches in the diagnosis of infectious disorders in brain or spinal cord biopsies in patients with suspected central nervous system (CNS) infections has been investigated. This NGS technology can provide a view of the transcriptome of the host tissue as well as capture microbial genomes such as bacteria, fungi, and viruses that reside in the tissue niche. Deep sequencing of total DNA or ribonucleic acid (RNA) provides an unbiased approach that can detect even rare components of the microbiome. A team of scientists at Johns Hopkins University (Baltimore, MD, USA; www.jhu.edu) performed a prospective pilot study, and applied NGS in combination with a new computational analysis pipeline to detect the presence of pathogenic microbes in brain or spinal cord biopsies from 10 patients with neurologic problems indicating possible infection, but for whom conventional clinical and microbiology studies yielded negative or inconclusive results. Fresh frozen tissues from eight cases were sequenced immediately after biopsy and two other samples were from paraffinprocessed tissues. The team found that direct DNA and RNA sequencing of brain tissue biopsies generated 8.3 million to

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29.1 million sequence reads per sample, which successfully identified with high confidence the infectious agent in three patients for whom validation techniques confirmed the pathogens identified by NGS. Although NGS was unable to identify with precision infectious agents in the remaining cases, it contributed to the understanding of neuropathological processes in five others, demonstrating the power of large-scale unbiased sequencing as a novel diagnostic tool. Clinical outcomes were consistent with the findings yielded by NGS on the presence or absence of an infectious pathogenic process in eight of 10 cases, and were noncontributory in the remaining two. Carlos Pardo-Villamizar, MD, an associate professor of neurology, a senior author of the study, said, “By incorporating modern genetic sequencing techniques into pathology diagnostics, we were able to investigate the potential presence of infection in 10 subjects and found appropriate explanations of clinical problems in eight out of 10 patient cases examined in this study. We hope to develop this technique further as a way to bring the diagnosis rate of inflammatory brain disorders and infections closer to 100% so we can treat patients more effectively.” The study was published on June 13, 2016, in the journal Neurology: Neuroimmunology & Neuroinflammation. LINKXPRESS COM

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Rapid TB Test Accuracy Compromised By Mycobacterial Diversity hroughout the world, Mycobacterium tuberculosis (Mtb) is responsible for the vast majority of tuberculosis (TB) cases; however, there are several other closely related mycobacterial species that cause TB, all part of the Mycobacterium tuberculosis complex (Mtbc). Diagnostics for rapid confirmation of positive liquid cultures presumptive of M. tuberculosis bacteria, based on the detection of the MPT64 antigen, and are being used in many TB diagnostic laboratories worldwide. The diagnostic performance of these tests in West Africa, where TB is uniquely caused by the geographically restricted Mycobacterium africanum (Maf 1 and 2) and M. tuberculosis lineages, has not been properly assessed. An international team of scientists led by those from the Medical Research Council (MRC) Unit, (Serrekunda, The Gambia; www.mrc.gm) collected sputum samples prospectively from individuals with suspected TB between April and October 2014 and were all initially screened for the presence of acid fast bacilli (AFB) by Auramine microscopy. They compared the abundance of mpt64 gene product in sputum samples of patients with untreated pulmonary TB caused by Maf 2, the strain common in The Gambia or Mtb. The scientists then prospectively analyzed culture isolates from 173 patients with one of the rap-

id tests, the BD MGIT TBc Identification test kit (Becton Dickinson Microbiology Systems, Sparks, MD USA; www.bd.com). They repeated the same analysis with all samples that had tested negative on day zero and a random set of those that had tested positive using the SD Bioline Ag MPT64 Rapid test (Standard Diagnostics, Yongin-si, Republic of Korea; www.standardia.com). They observed no significant difference between the two tests. Gene expression of mpt64 gene was performed using multiplex realtime polymerase chain reaction (qPCR). The team found that 150 of the samples tested positive on day zero (the day when mycobacterial growth in culture was first recorded), with 23 (13.2%) testing negative at this time point. The accuracy was much higher (over 90%) for the Mtb samples, compared with less than 80% for the Maf 2 samples. At Day 10, 84% of Maf 2 samples tested positive compared with 98% of Mtb samples. By Day 90, 98% of both Mtb and Maf 2 samples tested positive. Based on these results, 22% of Maf 2 patients, and 10% of Mtb patients would have been wrongly classified as having non-TB mycobacteria if the tests had not been repeated after day zero. At the end of the 10-day window recommended by the BD MGITTM TBc ID manufacturer, 16% of all Maf 2 samples remained negative, compared with only 2% of Mtb samples.

The authors indicated that MPT64 tests need to be cautiously used in settings where Maf 2 is common. However, they also recognize that given the relatively low cost, limited technical expertise and shorter turnaround time associated with using rapid speciation tests compared to alternative speciation methods, MPT64 rapid tests will likely remain one of the preferred options for timely diagnosis of suspected TB despite the possibility of false negative results, and suggest that a negative MPT64 result would require confirmation by an alternative method. The study was published on July 7, 2016, in the journal PLOS Neglected Tropical Diseases. Image: The Bioline TBAg MPT64 rapid immunochromatographic identification test for the M. tuberculosis complex (Photo courtesy of Standard Diagnostics).

Microneedle Sampling and Analysis System for Painless Drug Monitoring microneedle sampling and analysis system was shown to accurately measure levels of the drug vancomycin in the interstitial fluid with results comparable to those obtained by classical blood testing. Therapeutic drug monitoring typically requires blood drawn from patients in an uncomfortable and often painful fashion. An excellent example is monitoring of the drug vancomycin, which, due to the possibility of life-threatening

toxic side effects, may require three to four blood draws per day. As an alternative, investigators at the University of British Columbia (Vancouver, BC, Canada; www. ubc.ca) and their colleagues at the Paul Scherrer Institut (Villigen, Switzerland; www.psi.ch) developed a painless and minimally invasive method using hollow microneedles to extract extremely small volumes (less than one nanoliter) of interstitial fluid to measure drug concentrations. This approach

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was justified by the fact that plasma and interstitial fluid are very similar. This similarity exists because water, ions, and small solutes are continuously exchanged between plasma and interstitial fluids across the walls of capillaries. The new system consisted of a small, thin patch that was pressed against a patientâ&#x20AC;&#x2122;s arm during medical treatment and measured the level of the drug painlessly without drawing any blood. The tiny needle-like projection was less than half a millimeter long, resembled a hollow cone, and did not pierce the skin like a standard hypodermic needle. The inner lumen of the microneedle was functionalized to be used as a micro-reactor during sample collection to trap and bind target drug candidates during extraction, without requirements of sample transfer. An optofluidic device was integrated with this microneedle to rapidly quantify drugs with high sensitivity using a straightforward absorbance scheme. Vancomycin is currently detected by using volumes ranging between 50-100 microliters with a limit of detection of 1.35 micromoles. The microneedle-optofluidic biosensor system was shown to be capable of detecting vancomycin with a sample volume of 0.6 nanoliters and a limit of detection of less than 100 nanomoles. The microneedle technology is being commercialized by the spin-off company Microdermics Inc. (Vancouver, BC, Canada; www.microdermics. com). Details of the vancomycin sampling and analysis system were published in the July 6, 2016, online edition of the journal Scientific Reports. LabMedica International November/2016

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Early Detection Among Benefits of Skin Cancer Screening at Primary Care Visits ccording to new research, skin cancer screenings performed by primary care physicians (PCPs) during routine office visits improves detection of potentially deadly melanomas and find them in earlier stages. The results were presented at the 52nd annual American Society of Clinical Oncology (ASCO) annual meeting (June 3-7, 2016, Chicago, IL, USA; http://am.asco.org). “Our findings suggest that PCP screening is an effective way to improve early detection of melanoma, which could potentially save lives,” said the study’s lead author Laura Ferris, MD, PhD, associate professor, University of Pittsburgh School of Medicine (Pittsburgh, PA, USA; www. medschool.pitt.edu) of the University of Pittsburgh Schools of the Health Sciences (UPMC; Pittsburgh, PA, USA; www.upmc.com/pages/default.aspx) Skin cancer screenings are one of the most important steps for early detection and treatment. Typically, patients receive skin checks by appointment with a dermatologist. The goal of the new UPMC screening initiative, which was modeled after a promising German program, was to improve detection by making it easier for patients to get screened during rou-

tine office visits with their PCPs. PCPs completed training on how to recognize melanomas and were asked to offer annual screening during office visits to all patients aged 35 and older. In 2014, during the first year of the program, 15% of the 333,788 eligible UPMC patients were screened. On average, the melanomas detected in the group who received a screening at a PCP visit were nearly twice as thin as those detected in the group that was not screened by a PCP. Thinner melanomas have a better prognosis than more advanced thicker ones, so the new findings suggest PCP screening can find melanomas at an earlier, more treatable stage. In addition, only 5% of people in the screened group versus 20% of the unscreened group had melanomas of over 1 mm thickness, which are more likely to metastasize and require a biopsy of a nearby lymph node. “The PCP screenings prevented a lot of people from needing more aggressive therapy. Additionally, we did not see a high rate of false positive biopsies, in which no skin cancer was present, nor did we see a high rate of unnecessary dermatology referrals or skin surgeries, all of which suggest that the program did not simply drive up

health care costs needlessly,” said Dr. Ferris. Another important finding was that nearly half of the screened patients were men. Men are more likely to get and die from melanoma than women, but have been underrepresented in other skin cancer screenings published to date. “It’s exciting that our approach improves detection in this especially vulnerable population,” said Dr. Ferris. Image: The ability of cancer cells to move and spread depends on actin-rich core structures such as the podosomes (yellow) shown here in melanoma cells. Cell nuclei (blue), actin (red), and an actin regulator (green) are also shown (Photo courtesy of Julio C. Valencia and Daniel Mietchken, via Wikimedia).

Lab Offers New Generation Sequencing Test for Wound Microorganisms NA sequencing technology underlies a new diagnostic service that claims to identify all bacteria, fungi, and parasites as well as select viruses in a wound or infected area. The “Wound~Seq” test, which is performed on swab samples taken from the affected area, is available through Granger Diagnostics (Richmond, VA, USA; www.grangerdiagnostics.com). Samples collected by the doctor are sent to Granger’s laboratory, and results are usually returned within 48 hours. The Wound~Seq assay utilizes state-of-the-art

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DNA sequencing technology to identify all microorganisms present in the wound and does not rely on culture methods. It is the second NextGeneration Care Test in Granger Diagnostics’ pipeline, following Gynecologene, a test for bacterial vaginosis that incorporates this DNA sequencing technology. Dr. David G. Bostwick, chief medical officer at Granger Diagnostics, said, “We are excited to make available, for the first time, our clinicallysignificant test, Wound~Seq, that utilizes next-

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generation sequencing of the skin microbiome. Wounds and infections afflict millions of Americans every year, and are often perplexing to treat owing to the presence of unknown pathogens. Our new validated test provides significant actionable information that assists physicians in managing infections. Previous studies have shown that wound closure that relies on genetic-assisted testing is much more rapid, decreases pain and suffering, and results in cost savings. The promise of personalized medicine is now fulfilled.”

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Field Test for Zika Virus Combines PCR Components into a Single Low Cost Unit prototype PCR-based field test for the detection of the Zika virus (ZIKV) incorporates all required components and delivers accurate results in only about 40 minutes. Rapid and reliable diagnostics for ZIKV are vital because ZIKV-infected individuals display no symptoms or nonspecific symptoms similar to other viral infections. Since immunoassays lack adequate sensitivity and selectivity and are unable to identify active state of infection, molecular diagnostics are an effective means to detect ZIKV soon after infection and throughout pregnancy.

Investigators at the University of Pennsylvania (Philadelphia, USA; www.upenn.edu) recently described a highly sensitive reverse-transcription loopmediated, isothermal amplification (RT-LAMP) assay for rapid detection of ZIKV. The benefit of using the RT-LAMP procedure was that while it required that the sample be kept at a specific temperature, it did not demand that it be cycled through multiple precise temperature changes as in RT-PCR. On the other hand, RT-LAMP required more highly specialized DNA primers than did RT-PCR. To deal with the primer problem, the investigators identified highly conserved regions of the Zika virus genome that were divergent from other known pathogens. They then designed appropriate primers to recognize this sequence. The investigators then engineered a low-cost, point-of-care system that consisted of a diagnostic cassette and a processor. The cassette isolated, concentrated, and purified nucleic acids from a patient’s saliva sample and then carried out enzymatic amplification. The test results were indicated by the change in the color of a dye, which was inspected visually. For thermal control of the cassette, they used a chemically heated cup – adapted from military ration kits – without need for electrical power. The investigators demonstrated the utility of this novel POC diagnostic system by detecting ZIKV in oral samples with sensitivity of five plaque-forming units (PFU) in less than 40 minutes. This low cost technology (estimated to be about two USD per test device) is particularly suitable for resource-poor settings, where centralized laboratory facilities, funds, and trained personnel are in short supply, and for use in doctors’ offices, clinics, and at home. “Our work represents a proof of concept at this stage,” said contributing author Dr. Haim Bau, professor of mechanical engineering and applied mechanics at the University of Pennsylvania. “Before the assay can be adapted for medical use, we must experiment with patients’ samples and make assure that our assay and system match the performance of the gold standard and operate reproducibly and reliably. We are fortunate to have dedicated colleagues in endemic regions ready to assist us in this task.” The ZIKV field test was described in detail in the June 16, 2016, online edition of the journal Analytical Chemistry. Image: The prototype of a rapid, lowcost genetic test for the Zika virus. The patient provides a saliva sample, and the appearance of a blue color in the device’s lid indicates the presence of the virus (Photo courtesy of the University of Pennsylvania). LabMedica International November/2016

Edited by Tahir Pillay MBChB, PhD, FRCPath(Lon), FCPath(SA) IFCC members may send news to: Tahir Pillay MBChB, PhD, Head, Dept of Chemical Pathology, Faculty of Health Sciences, University of Pretoria, Private Bag Bag x323, Arcadia, 0007, South Africa Tel: (27) 012-319-2114; Fax: (27) 012-328-3600; Email: enews@ifcc.org

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Prof. Howard Morris of Australia Elected IFCC President for 2018-2020 To Serve as President-Elect in 2017 he IFCC is pleased to announce that Prof. Howard Morris (Adelaide, Australia), has been elected as IFCC President elect. The term of IFCC President elect will commence on January 1st, 2017, to be confirmed as IFCC President for the period January 1st, 2018 until 31st December 2020. We congratulate Prof. Howard Morris, and wish him a fruitful co-operation for the promotion of Clinical Chemistry and Laboratory Medicine world-wide. Prof. Howard Morris currently hold positions as Professor of Medical Sciences, University of South Australia, conducting research and teaching Laboratory Medicine and Clinical Scientist for SA Pathology, the public pathology provider for the state of South Australia. As a Clinical Scientist, he provides consultancy to medical practitioners and patients. With qualifications BSc(Hons), PhD, Fellow of the Australasian Association of Clinical Biochemists (FAACB) and Fellow of the Faculty of Science of the Royal College of Pathologists of Australasia (FFSc(RCPA), he has 40 years’ experience in Laboratory Medicine. His working experience includes provision of routine services, management of an endocrine laboratory, research administration with 6 years as Director of a medical research institute. His research interests include the pathophysiology of metabolic bone disease, and biomarkers of disease including bone turnover from which he has produced 280 peer-reviewed publications, achieving some 10,000 citations, with over 4,000 citations since 2011. AU$ 9 million has been awarded to fund his research. Prof. Morris’ professional activities include serving as IFCC Vice-President (2012-2014) contributing to policy development for our profession and the IFCC to meet the requirements of the 21st century. He has previously served on the Executive Committee of the Scientific Division of the IFCC including Secretary, held memberships of two IFCC Task Forces, Global Strategy for Diabetes Mellitus and Clinical Liaisons, and International Scientific Committee, XXI WorldLab, Berlin, 2011. A key aspect of his IFCC work has been to enhance interaction between Laboratory Medicine and clinical disciplines and patient groups. Within the Asia Pacific Federation of Clinical Biochemistry and Laboratory Medicine (APFCB) Prof. Morris served as Chair, Scientific Committee (2002 to 2005) and member Organising Committee, 10th Asia Pacific Congress of Clinical Biochemistry. Nationally he has served the AACB for over 30 years as Branch Chair and Education Representative, Chair or member of the Scientific Committee for AACB National Scientific Meetings and

IFCC OFFICE Via Carlo Farini 81, 20159 Milan, ITALY Tel: (39) 02-6680-9912 • Fax: (39) 02-6078-1846 E-mail: ifcc@ifcc.org • Web: www.ifcc.org Office Hours: 8.30-13.00 and 13.30-17.30 Staff Members: Paola Bramati, Silvia Cardinale, Silvia Colli-Lanzi

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Convenor of Workshops, Chemical Pathology Courses and Symposia. Between 1998 and 2004, Prof. Morris served on the AACB Federal Council as National Representative to IFCC and APFCB and Editor of the Clinical Biochemist Reviews, published by the AACB in conjunction with the APFCB (1994 to 2002). In 1993 he was an AACB Current Concepts Lecturer; in 2003 he was awarded an AACB Outstanding Service Medallion, 2004 the AACB W Roman Travelling Lectureship, 2009 Bio Innovation SA Bioscience Industry Leader Award, Louis Avioli Mecont’d on page 58

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Prof. Howard Morris of Australia Elected IFCC President for 2018-2020 cont’d from page 57

morial Plenary Lecturer, American Society of Bone and Mineral Society, Finalist Excellence in Research for Public Good, South Australian Science Excellence Award, 2012 Association of Clinical Biochemists (United Kingdom) International Lecturer, 2015 Australian and New Zealand Bone and Mineral Society Career Achievement Award.

Prof. Howard Morris Position Statement Laboratory medicine faces significant challenges and the IFCC requires a dynamic Executive Board to provide leadership for our profession. These challenges face all in the delivery of healthcare however Laboratory Medicine faces particular issues arising from its perception as merely a service provider rather than a driver for optimal healthcare. Clinical laboratories are required to assume increased responsibilities for service provision and patient safety with ever decreasing budgets. Financial constraints also impact our corporate members, moderating their ability to collaborate in professional development and, at the same time, our academic basis is diminishing. Therefore IFCC professional leadership has never been more vital to provide strategies for improving the crucial role of Laboratory Medicine in patient care. To build on the foundation of 60 years of IFCC leadership in Laboratory Medicine I believe the following areas are critical. (1) Improving clinical laboratory performance through standardization and traceability as it impacts on the rational use of clinical laboratory resources. Interna-

tional harmonization activities will further contribute to this field during the next triennium and the IFCC is required as a leading partner. A major challenge is to extend these activities to regions in which the current multi-national In-Vitro Diagnostic industry members are not the major provider of clinical laboratory instruments and reagents, in particular in the Asia Pacific as well as Central and South American regions. (2) Continue educational and training activities amongst our membership capable of meeting the needs of both developing and developed communities across general and specialised services including promotion of eLearning opportunities. (3) To ensure the financial sustainability of the IFCC (4) Promote closer interaction between international organizations representing various disciplines of Laboratory Medicine to provide common engagement by our profession with clinical professionals and their international organizations as well as government organizations and other stakeholders in healthcare. (5) Enhance communication between our members through electronic platforms and increasing availability in non-English languages. (6) Extending our partnerships to relevant patient organizations. The IFCC is a federation of national societies and corporate members. The major resource of the IFCC is the voluntary contributions of individual professionals arising from our membership. Thus the effectiveness of the IFCC is dependent on this committed support of

members to meet our goals and commitments. The next EB is required to enhance and strengthen the relationships upon which our organization is founded. As President I will work within the EB to direct the IFCC Divisions, Task Forces, Committees and Working Groups to meet strategic goals ensuring the widest consultation and discussion amongst the National Society representatives, the regional federations and our members as we chart our progress for the future development of Laboratory Medicine. My current employers fully support my professional activities and should I be elected I will be able to devote up to 75% of my time to IFCC responsibilities. As a Visiting Lecturer for IFCC I have discussed issues facing our profession with national representatives of 24 IFCC members and, as well, presented at 23 international or regional congresses. I look forward to continuing these discussions, for you to provide your suggestions as to how best IFCC can assist to improve the practice of Laboratory Medicine in your country. Howard Morris PhD FAACB FFSc(RCPA) Participation in the new electronic voting format has been very high. The elections have been conducted via electronic system in order to ensure the wider participation to this important moment in the IFCC life. Results can be seen at www.ifcc.org The call for nomination for IFCC Secretary and IFCC Treasurer is currently open, until 15th December 2016. The Executive Board Members (Regional Members) election will take place in 2017.

Jordanian Society Launches National Magazine by Lina Mashhour Assaf, BSc., MSc., Medical Laboratory Professional, Chief Editor of Lab Pulse Magazine

Photo: Executive board of the Society with official sponsor the deputy of Minister of Health.

n 23 July 2016, Medical Technology and Laboratory Society, Jordan, launched the first issue of its periodic magazine “Nabd Al-Mukhtabar” (Laboratory Pulse) during a celebration and dinner party carried out under the patronage of his Excellency the Minister of Health. The magazine will be published three times a year and will specialize in health areas, particularly in the field of medical technology and laboratory medicine hence the name “Lab Pulse”. It will serve the medical laboratory industry and those interested in this area from other health disciplines, in addition to the general public. The main goal of creating this magazine was to serve laboratory personnel, through the transfer and exchange of experiences and providing updates of new developments in the different areas of specialization, and to increase the awareness of the importance of their work. In addition, it is meant to

strengthen the social and professional ties between them. Other objectives of creating this magazine were to contribute in the dissemination of health awareness among the general public, to educate them about the role of medical laboratory sector and shed light on the importance of this sector in the overall health process. The magazine also aims to communicate between medical and laboratory personnel where medical personnel can learn about the many modern and sophisticated laboratory analysis techniques, and about the extent of the efforts made by the laboratory professionals to obtain results with high levels of accuracy and precision, leading to a deeper understanding and greater respect of the role of medical laboratory staff, and thus to place the profession where it fits side by side among the other medical specialties. LabMedica International November/2016

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Finnish Society Celebrates 70 years by Heidi Nousiainen; Member of the FSCC Board he Finnish Society of Clinical Chemistry celebrated its 70th anniversary in April 2016. The festivities were held in the jugend-style design hotel GLO Hotel Art in Helsinki, Finland in conjunction with the biannual national meeting. The Finnish Society of Clinical Chemistry (FSCC) was founded in 1946 and was the first national society in Europe to incorporate the words “Clinical Chemistry” in its name. The purpose of FSCC is to act as a link between physicians and chemists with an interest in clinical chemistry and to promote the theoretical and practical development of clinical chemistry in Finland. The two-day celebratory meeting covered some of the hottest current topics in the industry, including an introduction into service design by the Finnish company Hellon, and a one-day LEAN workshop,

where participants could experience the power of LEAN hands-on by passing tennis balls to each other. The meeting also included a visit to the Helsinki University Hospital Laboratory HUSLAB’s brand new laboratory building that hosts the longest automation track in Europe as well as cutting-edge facilities specifically designed to meet the needs of modern lab technology. The festivities culminated in a celebratory dinner, with former FSCC presidents invited as honored guests. Almost one hundred society members enjoyed good food, excellent company, and entertainment provided by standup-comedian Mikko Vaismaa. The night ended with live music provided by the talented musicians of Flipper orchestra. The FSCC actively continues to promote clinical chemistry in Finland. The next big event organized by

FSCC is the 36th Nordic Congress for Clinical Chemistry which will be held in Helsinki on June 12th-15th, 2018. We look forward to meeting you there!

IFCC-EMD Working Group On Flow Cytometry: A Report by Prof. Ulrich Sack Krems, Austria. “Extracellular Vesicles in Inflammation“ have been the topic of a meeting. Beside cutting edge presentations covering this topic, practical exercises were offered to the attendees. Furthermore, the flow cytometry workshop took place in Athens, Greece. The established principle was continued. The first Latin American Course was offered in Cordoba/Argentina in 2016. Local support was given by the Rotary Club de Sierras Chicas and by the University of Cordoba. Course participants get personal information folders with all presentations and additional material. Another flow cytometry workshop will take place from October 26 to 28 in St Petersburg (Russia). The established principle will be continued: intense interaction between participants and trainers; 3 topics a day with short introduction and extensive practical exercises; 3 day duration; and emerging and relevant scientific topics with clinical impact for daily practice. For 2017, further courses in Latin America and Europe are in preparation. In February, an additional flow cytometry winter school will be offered in SaintEtienne, France. We will continue the successful principle of our courses: Intense interaction between participants and trainers; 3 topics a day with short introduction and extensive practical exercises; 3 day duration; Personalized information folders with all presentations and material for practice at home; Emerging and relevant scientific topics with clinical impact for daily practice. Last not least, the 2009 edition of Cellular Diagnostics by Sack, Tarnok, and Rothe (Karger publishers) is still the most recent book in clinical flow cytometry and has been supported by the IFCC.

low cytometry working group focusses on promoting recent developments in clinical flow cytometry. This is done in co-operation with national and international scientific organizations. Main activity is organizing courses. The scientific organizer is U Sack (Leipzig, DE), supported by C Lambert (St. Etienne, FR), A Spittler (Wien, AU), K Psarra (Athens, GR), C Rodriguez (Cordoba, AR), and AM Ivanov (St Petersburg, RU). Technical support, organization and technical equipment is generously provided by Beckman-Coulter. The courses have a long tradition. Following an intermediate disruption in 2007, first new flow cytometry course took place in 2012 in Leipzig, Saxony, Germany. The focus was flow cytometry in immunology and stem cell research. This course was given by specialists in analytic cytometry mainly outside hematology but including sorting. Local scientific supporter was the Translational Centre for Regenerative Medicine Leipzig (http://www.trm.unileipzig.de). Feedback by participants indicated strong appreciation with the high percentage of practical exercises. Second course was organized in 2013 in co-operation with the ESCCA in Saint-Etienne, France. The focus was the monocyte-macrophage system. Parts of this course have been published (Monocytes and macrophages in flow, Lambert et al., Cytometry B). Again, the hands-on time and the scientific level were scored excellent by the participants. The third course in 2014 was locally organized by Andreas Spittler in Vienna, Austria, again in co-operation with the ESCCA. Participants enjoyed the closed contact with experienced flow cytometrists and the intense hands-on training. In 2015, an additional workshop was offered under IFCC auspices in

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Developing Quality Competence in Medical Laboratories: A Historical Perspective by Dr Michael Thomas, Chair, Special Project for Developing Quality Competence in Medical laboratories s my term of office as Chair of the Special Project for Developing Quality Competence in Medical Laboratories (DQCML) comes to a close at the end of this year it is perhaps appropriate to summarise its origin, ambitions and accomplishments since it was established a decade ago. It was at the IFCC Executive Board meeting in Asuncion, Paraguay in 2006 under the presidency of Professor Hicks that there was recognition by the Board that the extent and quality of laboratory testing in many developing countries lagged behind what is commonly accepted as needed in advanced countries. In many developed countries it is often stated, although with limited scientific proof, that as much as 80% of medical decisions are based on laboratory test results. This underlying importance of laboratory testing is a primary reason why IFCC, within the limits of its resources, determined to assist the progressive evolution of laboratory testing in developing countries. The Executive Board challenged the Education and Management Division (EMD), at that time under the leadership of Janet Smith, as to ways in which support could be provided to improve the quality process within medical laboratories. EMD’s response was to propose a Special Project be established aimed at informing emerging laboratory services through their National Societies of the opportunities offered by IFCC to

develop local quality competence which would focus on all aspects of quality, but concentrate particularly on internal quality control, external quality assessment and working towards laboratory accreditation with the aim of adoption of a quality system in line with the international standard ISO 15189. EB approved these recommendations and this led to the setting up of an Oversight Board under the direction of EMD that subsequently became known as the Special Project Group for Developing Quality Competence in Medical Laboratories (DQCML) in March 2007. The Inaugural meeting of the Special Project Group took place in Amsterdam in June 2007 where it was immediately recognised that many of the aims and objectives of the project as proposed in a Quality Ladder starting at Internal Quality Control and culminating after a number of steps in ISO 15189 Accreditation would take a long time to achieve and it seemed sensible to consider the adoption of a modular approach. The basic objective for the Project was agreed as being to assist in improving the quality of a laboratory medicine service as this was seen as an essential requirement for better patient care and improved clinical outcomes. It was of course recognised that this was no mean task and that to achieve it would take many years. Members who would likely seek advice from the Special Project would probably be at different stages of their

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development in implementing a Quality System. It therefore became convenient to consider the process as a series of steps on a Quality Ladder with the possibility of joining the ladder at any point. Furthermore it was recognised that the Quality Ladder would itself conveniently split into two components Firstly for those countries that would benefit from advice on the introduction of quality control and external systems of quality assurance And secondly for those countries that had a more mature understanding of quality and would benefit from more advanced help in working towards accreditation. Funding was made available to provide advice and assistance rather than any on-going commitment to maintain systems already in place. And, at the outset, Professor Hicks had secured additional funding from Abbott Diagnostics for the Visiting Lecturer Programme and the DQCML would have access to that funding source Throughout its lifetime, the DQCML Project has drawn hugely on the experience and expertise of the Committees for Analytical Quality and Clinical Laboratory Management and more recently the Committee for Distance Learning. It has been through colleagues on all three of these committees that it has been possible to offer support workshops, lectures and webinars geared to the particular rungs of the Quality Ladder template. The first initiative delivered jointly by DQCML and the APFCB was a 5day workshop in Spring 2009 in Sri Lanka on the “Role of accreditation in assuring analytical quality”. The workshop was expertly led by Sam Vasikaran on behalf of APFCB and by Janet Smith on behalf of EMD and the DQCML and in keeping with the aims of the DQCML of developing transferable educational modules was later repeated in Macedonia. The second dovetailed with a project in Vietnam being undertaken by the AACB under Renze Bais and which supported the Vietnamese in developing an internal programme of EQA. Subsequently the DQCML activities have impacted on many member countries across the globe including Indonesia, Tunisia, The Philippines, Uruguay, Nepal, Romania and Russia. Most recently the project has supported the delivery of two workshops in Ecuador. These workshops were delivered in Spanish which limits them being shared more widely amongst the IFCC membership but recognises the significant Spanish-speaking representation within IFCC. Over the ten years of its existence, the Project has benefitted from the

huge expertise of its colleagues within EMD. The achievements of the DQCML has depended and ultimately succeeded through the collaboration, assistance and expertise of colleagues on other committees, notably that from C-AQ, CCLM and C-DL. The educational programmes the DQCML have delivered have been bespoke to the applicant member. Hence although they could be useful to other members there is, as yet, no consolidated resource that can be accessed. However it still remains the ambition and the ultimate aim of this Special Project to publish a bespoke Quality Manual bringing together the various initiatives and reflecting the stages in developing Quality Systems as a progression through each rung of the Quality Ladder. More recently the DQCML has been encouraged to consider an extension of its role into Sub-Saharan Africa where it will need to consider the Stepwise Laboratory Quality Improvement Process Towards Accreditation or SLIPTA way of achieving accreditation. Our first steps in that area will be in delivering through a one-day symposium and workshop in October 2016 at the 6th Scientific Conference of the Association of Clinical Chemists of Nigeria entitled “Sustaining Quality Practice and Process in Developing Countries”. This opens a new and exciting door on DQCML activities. It has meant that with the support of IFCC Presidents past and present that the DQCML Special Project continues and expands with renewed opportunities, priorities and challenges and it sees the Quality Ladder evolving with significantly greater challenges and responsibilities than originally imagined. Over the last ten years I believe that the DQCML has made a significant and useful impact in helping IFCC Members improve the quality of their laboratory medicine services, I wish my successor, Professor Egon Amman every success in leading the DQCML towards enhancing the quality of laboratory medicine services across the globe. LabMedica International November/2016

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EFLM CORNER

European Federation of Clinical Chemistry and Laboratory Medicine

Editor: Harjit Pal Bhattoa, MD, PhD, EuSpLM

FOREWORD

Dr Harjit Pal Bhattoa, Editor

his current issue of the EFLM New Corner summarizes the pleasant experiences of two EFLM bursary recipients, Steven De Keukeleire and Helin Tuukka, at the 4th Joint EFLM-UEMS Congress “Laboratory Medicine at the Clin-

ical Interface” held in Warsaw, Poland. MariaStella Graziani, Chair of the EFLM Communication Committee presents an update to the list of 2016 EFLM publications. The EFLM Education and Training Committee present’s a short summary of their activities. Daniel Rajdl, Chair of the EFLM WG Distance Education and eLearning, introduces the upcoming webinars.

4th Joint EFLM-UEMS Congress “Laboratory Medicine at the Clinical Interface”: Reports from Bursary Recipients Helin Tuukka, EFLM Bursary Recipient First, I would like to express my sincere gratitude to the EFLM, giving me the opportunity to attend this congress. Generally, the congress was well organized with a wide variety of topics. The magnificent opening lecture by Prof. Dennis Lo, deserves a special mention as well as the lively debate on vitamin D measurement (Pro: Roger Bouillon, Contra, Ian Young) and the practical and important EFLM-EAS consensus recommendation on nonfasting dyslipidemia testing, illustrated by Michel Langlois. I presented two posters at the congress and received positive feedback. I had the honor to present one of them as part of a poster walk session. Since my research field is coagulation, I was interested in the debate on

monitoring direct oral anticoagulants (DOAC), with Prof. Lotta Joutsi-Korhonen from Finland taking the prostance and Prof. Grzegorz Grześk from Poland taking the contra-stance. Dr Joutsi-Korhonen started her presentation by comparing the DOAC with warfarin. The INR is used for warfarin regular monitoring due to the narrow therapeutic range. With the direct oral anticoagulants too, while routine coagulation monitoring is currently not recommended, there are a number of clinical situations where the monitoring is mandatory. In case of complications with DOAC treatment, which are numerous (treatment failure, trauma, emergency procedure, bleeding or overdose), it is critical to know the DOAC level. Coagulation screening tests PT and APTT are not suitable, but thrombin time and anti-Xa assays

should be used. Finally, there is a consensus that blood count, liver and renal function should always be monitored. Prof. Grześk started his presentation on INR: since warfarin has a narrow therapeutic range and the monitoring is mandatory, we think that also DOAC need monitoring. However, the differences are numerous; the proper dose is usually prescribed without any laboratory tests and indeed, there is no recommendation regarding routine monitoring. When patients are asked what they expect to ensure safety, the most frequent response is presence of an antidote – i.e. they do not feel safe because of laboratory monitoring of drug effect. Furthermore, in the routine practice very few requests about DOAC are observed; in prof Grześk laboratory, in the year 2016 only two requests for dabigatran concentration have been received. At the end of the debate, the chair suggested a consensus: patients should be always monitored to assess liver and renal function, with coagulation monitoring in selected cases, not routinely. Both speakers agreed on this. I’m quite partial in my assessment, since Lotta Joutsi-Korhonen is my supervisor, but I think that there are many arguments in favor of DOAC monitoring and that her thesis emerged as winner in this debate.

Steven De Keukeleire, EFLM Bursary Recipient The 4th joint EFLM-UEMS Congress was held from the 21nd to 24nd of September 2016 in Warsaw, Poland. The conference was organized under the IFCC auspices, the honorary patronage of the Mayor of Warsaw, the Medical University of Warsaw and Collegium Medicum of the Nicolaus Copernicus University. The congress was attended by more than 450 scientists and physicians. The programme covered a large range of clinically important emerging topics in laboratory Medicine focusing on the Clinical Interface between the laboratory and clinical practice. Hot topics about cardiovascular disease, diabetic kidney disease, therapeutic drug monitoring, pediatric laboratory medicine, auto immunology, biomarkers and molecular diagnostics were discussed. One session was dedicated to “Trends in pediatric laboratory medicine” and included presentations about pediatric obesity, newborn screening,

pediatric reference intervals related to therapeutic drug monitoring (TDM) of antibiotics. Childhood obesity is a worldwide problem that is difficult to manage. It often results in diabetes. There is a need for adequate laboratory assessment by measuring cholesterol (total cholesterol and LDL-levels), glucose (using Hb A1C concentrations), and liver parameters as well (transaminases). Growing evidence points to a role of the intestine in obesity. Newborn screening (NBS) for lysomal diseases (Fabry, Pompe, Gaucher, MPS type I) has gained interest due to the increasing number of therapeutic options. The analytical performances of the laboratory methods are essential to improve the clinical outcome. Despite the great improvement, concerns remain about the appropriateness of screening methods, the effectiveness of therapies and the overall costs involved. To define pediatric reference intervals a great initiative was the CALIPER (Canadian Laboratory Initiative on Paediatric Reference Intervals) project resulting in a usable database with normal reference values for a wide range of biochemical markers between 0 and 18 years of age. Another topic discussed was the antibiotic Meropenem usage in the neonatal population. There is a big need for adequate TDM in newborns. The biochemical stability of Meropenem in biological samples is crucial since the measurement is performed in specialised laboratories and there is the need for iced transportation and long term storage at -80°C. The diversity of scientific topics, the excellent organization and social programme in Warsaw fully contributed to a successful 4th Joint EFLM-UEMS Congress. Young scientist were rewarded several grants by the EFLM to attend the congress. EFLM stimulates membership of young scientists offering a bursaries programme, and establishing educational projects in order to increase the interest in laboratory medicine throughout Europe. Recognition of specialists in laboratory medicine remains an important issue. Individuals who practice with knowledge, skills and competence at an (inter)national level can make the difference. Based on this experience, joining the EFLM- community offers a multitude of possibilities. LabMedica International November/2016

EFLM CORNER

European Federation of Clinical Chemistry and Laboratory Medicine

EFLM Publications in 2016: An Update by Maria Stella Graziani, Chair of the Communications Committee hree more papers have been published by EFLM functional units since the last issue; please find below the list. The papers are freely downloadable at the dedicated page of the EFLM website (www.eflm.eu/index.php/ eflm-publications.html)

been written for laboratories and the ones that affect laboratories are inadequate: we need to change this culture and ensure that we are involved in both the arenas of diagnostic research and guideline writing.

Reviews & Surveys Positions & Opinion Papers Barth JH, Misra S, Aakre KM, Langlois MR, Watine J, Twomey PJ et al. by the EFLM-UEMS joint WG on Guidelines. Why are clinical practice guidelines not followed? Clin Chem Lab Med 2016 DOI 10.1515/cclm2015-0871 An interesting opinion paper that investigates the reasons why clinical practice guidelines are so poorly followed both by clinicians and by laboratory medicine. Few guidelines have

Ceriotti F, Gligorovic Barhanovic N, Kostovska I, Karel Kotaska K, Perich Alsina MC , on behalf of the EFLM Working Group on Harmonisation of total testing process. Harmonisation of the laboratory testing process: need for a coordinated approach. Clin Chem Lab Med 2016 DOI 10.1515/cclm-2016-0244 A survey aimed to collect information on the harmonisation activities by the different national societies member of EFLM. The results of the survey indi-

News from the EFLM Education and Training Committee

cate that there are some harmonisation initiatives in place in Europe, but these initiatives are not coordinated. Considering that the analytical phase is already covered by specific projects, EFLM is focusing its harmonisation efforts on the pre- and post-analytical phases. Plebani M, O’Kane M, Vermeersch P, Cadamuro J, Oosterhuis W, Sciacovelli L on behalf of the EFLM Task Force on “Performance specifications for the extra-analytical phases” (TFGPSEP). The use of extra-analytical phase quality indicators by clinical laboratories: the results of an international survey. Clin Chem Lab Med 2016 DOI 10.1515/cclm-2016-0770 The paper reports about the first initiative of the Task Force: a questionnaire administered to all National Soci-

eties of the Federation and other stakeholders aimed to understand the stateof-the-art on Quality Indicators (QIs). The data confirm the existence of the QI paradox as all responders were aware of the need to implement QIs and related performance criteria in their laboratories but the number and type of QIs monitored varied significantly. There is an important role for national societies and international federations to increase awareness in clinical laboratories and to encourage participation in initiatives to develop consensus on the QIs to be employed and the related performance criteria. The first initiative of the Task Force was to understand the state-of-the-art on QIs using a questionnaire administered to all National Societies of the Federation and other stakeholders.

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head of the 4th Joint EFLM-UEMS Congress the members of the EFLM Education and Training Committee (C-ET) discussed different ongoing projects together with their Chair Ralf Lichtinghagen. Recently the Task Finish Group Continuous Professional Development (TFG-CPD) started with Elizabeta Topic (chair) and additional 17 experts from the different EFLM member countries it work with. The main tasks of the group are on one hand the preparation of guidelines for the accreditation of CPD events and on the other hand the guidelines for certification of European Specialists in Laboratory Medicine. An action plan suggested from the chair was discussed, initial results are now expected at the end of this year. All members of the C-ET verified during the session that the generation of an EFLM CPD crediting system will be an important step especially for the whole group of non-medical laboratory specialists who often have no access to any kind of existing (e.g. national) crediting system.

LabMedica International November/2016

The Working Group Congresses and Postgraduate Education (WGCPE) presented under the chair Gustav Kovac different ongoing projects. Of main importance the group suggested the final development of an Educational and Training Exchange Program for laboratory professionals, which was exemplary initiated by the WG group member Evgenjia Homsak. The EFLM Executive Board just endorsed this project, the committee members emphasized that they await optimistically the project approval by the EFLM EB at the end of the year. Daniel Rajdl, chair of the Working Group Distance Education (WG-DE), reported about his evaluation of the last webinars, which were successful events. Recordings of the sessions were, especially on YouTube, downloaded from interested persons all over the world. Actually the WG is looking forward the recording of selected sessions of the 4th EFLM-UEMS Congress to present it later for educational purposes.

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EFLM CORNER Upcoming Webinars Daniel Rajdl, Chair, EFLM WG Distance Education and e-Learning aniel Rajdl, Chair of the EFLM WG Distance Education and e-Learning reports about next webinars. EFLM is pleased to remind you that the attendance to the webinars is free of charge and that the recording of the lectures will be available later on at the EFLM web site for those unable to attend.

Biomarkers in Guiding Treatment of Heart Failure Nov 17, 2016; 18:00 CET - Speaker: Alan S. Maisel; Moderator: Evgenija Homsak About the speaker: Alan S. Maisel is Professor of Medicine at the University of California, San Diego (UCSD) and Director of the Coronary Care Unit and

European Federation of Clinical Chemistry and Laboratory Medicine Heart Failure Program at the VA San Diego Healthcare System in La Jolla, California. Prof Maisel is active on the faculty at UCSD where he won numerous teaching awards; he has just completed a ten-year stint as Associate Editor of the Journal of the American College Cardiology. Prof Maisel is considered one of the globally recognized experts on cardiac biomarkers and has over 400 scientific publications. He has authored several ground-breaking manuscripts that have paved the way for development of diagnostic tools for patients with congestive heart failure. In particular, he was the leading investigator on studies that brought the use of BNP into clinical practice and was the lead investigator on seven multicentre biomarker trials. Prof Maisel is actually working to delineate the clinical role of sST2 levels in clinical practice for guiding treatment. Summary. Heart failure represents an important clinical problem and the accuracy of diagnosis by clinical means alone is often inadequate, especially in the early, asymptomatic stages of the disease. For these reasons, there is an increasing interest in the development of cardiovascular biomarkers useful for diagnosis, prognosis, follow-up of patients with heart failure and to possibly guide treatment as well. In this talk, three biomarkers will be examined: natriuretic peptides, high sensitivity troponins and sST2. Natriuretic peptides (NPs) (BNP and NTproBNP) are well recognized biomarkers for the diagnosis of heart failure and have been included in clinical guidelines. The limitations for their use include the wide variability of their concentration within subjects and the possible non-specific increase. Additionally, their clinical value is questionable in patients receiving Entresto (a drug that inhibits breakdown of NPs). High sensitivity troponins in the setting of acute heart failure (and maybe chronic) are a marker of subendocardial necrosis and may indicate a worse prognosis. It can be postulated that drugs like nitrates could be used in heart failure treatment more efficiently when levels of high sensitivity troponin are high. Finally, sST2 is a marker of fibrosis and is elevated in virtually all patients with heart failure. In the acute setting, it defines a subset of patients in need of advanced treatment to avoid rehospitalization. In the chronic setting, titrating treatment to a sST2 level below 35 ng/ml appears to mitigate the risk, even if NPs are not decreasing. The data from “Entresto” clinical trial seem to suggest that sST2 levels are useful both to select patients eligible for the treatment with the drug and to dose it and for monitoring purposes as well. Save the Date. Next scheduled EFLM webinars:

Case report: Patient with shock and multiorgan failure Dec 13, 2016; 18:00 CET Speaker: Anna Merino

Reliable estimates of biological variation – the way forward May 9, 2017; 18:00 CET Speaker: Aasne Karine Aarsand LabMedica International November/2016

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Industry News

International Calendar

Beckman Parent to Acquire Lab Consumables Producer anaher Corporation (Washington, D.C., USA; www. danaher.com), parent of Beckman Coulter, Inc., has agreed to acquire Phenomenex Inc., (Torrance, CA, USA; www.phenomenex.com), a privately held manufacturer and distributor of high-value consumables for the separation sciences, including proprietary chromatography consumables for the liquid and gas chromatography market. Phenomenex has more than 7,000 products supporting various applications in drug discovery and development, clinical research, forensic toxicology, petrochemical, environmental analysis and food safety. Following the acquisition, it will operate as a stand-

alone operating company, retaining the Phenomenex brand, its personnel and site locations. “Phenomenex is a widely recognizable brand that we are proud to say is highly respected and well established as providing high-quality consumables to the scientific community worldwide,” said Fasha Mahjoor, CEO, Phenomenex. “Joining Danaher will allow us to maintain the high pace of innovation that our customers and international distributors have come to expect from us. This is our opportunity to further expand our R&D activities, benefit our customers with a more diversified product portfolio and reinforce our market leadership.”

Middle East IVD Market To Reach USD3.4 Billion by 2021 he Middle East market for in vitro diagnostics will be worth USD 3.4 billion by 2021 and record a growth that is one and one-half to double that of the world IVD market. The growth will be driven by expanding populations and a commitment to healthcare spending in this region, which houses offices of most global concerns in Saudi Arabia, Egypt, Dubai or other locales to service this population with testing products and supplies. These are the latest findings of Kalorama Information, (New York, NY, USA; www.kaloramainformation. com), an independent medical market research firm.

In the 1970s, a number of Middle East and Gulf countries began providing public healthcare to their citizens. Currently, Oman, Saudi Arabia, Qatar, Syria and Turkey all have universal healthcare programs. Lifestyle changes in the Middle East, such as a more sedentary lifestyle that is resulting in rising obesity rates, has led to the emergence of “Western” diseases such as diabetes. This has resulted in a growing demand for medical tests in the region, thus necessitating increased investment in healthcare, which generally takes the form of government tenders that remain a major source of supply for state-controlled labs and hospitals.

Founder of Quantimetrix Dies at 89 r. Robert William Ban, who founded Quantimetrix Corporation (Redondo Beach, CA, USA; www.quantimetrix.com) over 40 years ago, and led the company's growth into a powerhouse in laboratory liquid control systems, died peacefully in California at the age of 89. Dr. Ban, a PhD in Bichemistry, started Quantimetrix in 1974 as a result a number of successful experiements he conducted in his garage. The company pioneered a liquid stabilization technology that simplified quality control methods in the laboratory. With the Clinical Laboratory Improvement Act (CLIA) of 1988 mandating the use of controls in clinical laboratories, Quantimetrix provided the products to a diagnostic community eager to abide by the new requirements. The demand for Quantimetrix products soared and established the company as a global

leader in the field. Earlier, in 1972, Dr. Ban also founded Diagnostic Products Corporation (DPC), a developer of radioimmunoassay (RIA) diagnostic test kits. The test, considered a breakthrough at the time, permitted the measurement of infinitesimally low concentrations of substances in bodily fluids, such as drugs and hormones. Dr. Ban left DPC in 1973 and subsequently started Quantimetrix. Dr. Ban was a storied innovator and a tireless leader who was active around the world. His son, Monty Scott Ban, continues to pursue his vision as the current chairman and CEO of the company. Besides his active business life, Dr. Ban was also a published author, an artist, poet, and musician. He is survived by his son, daughter-in-law, and three granddaughters.

For a free listing of your event, or a paid advertisement in this section, contact:

International Calendar, LabMedica International P.O.Box 802214, Miami, FL 33280-2214, USA Fax: 1-954-893-0038 • E-mail: info@globetech.net

NOVEMBER 2016 EFLM Course on Test Evaluation, Developing medical tests that improve patient outcomes. Nov 9-11; Leiden, The Netherlands; Web: www.eflm-test-evaluation-course.eu Association for Molecular Pathology (AMP) Annual Meeting 2016. Nov 10-12; Charlotte, NC, USA; Web: www.amp.org MEDICA 2016. Nov 16-19; Dusseldorf, Germany; Web: www.medica.de 10th International Scientific Meeting of the Centre of Metrological Traceability in Laboratory Medicine (CIRME) “Ten Years After”. Nov 17-18; Milan, Italy; Web: http:// users.unimi.it/cirme/home WASPaLM 2016- 29th World Congress of Pathology. Nov 18-21; Las Vegas, NV, USA; Web: www.waspalm.org 14th Asia-Pacific Federation for Clinical Biochemistry and Laboratory Medicine Congress. Nov 26-29; Web: www. apfcbcongress2016.org

DECEMBER 2016 50e Journées de Biologie Praticienne. Dec 2-3; Paris, France; Web: http://eflm.eu

JANUARY 2017 The British Fertility Society (BFS) Annual Meeting. Jan 5-7; Edinburgh, UK; Web: http://fertilityconference.org CBRD 2017 - 4th Caribbean Biomedical Research Days. Jan 16-18; Gros Islet, Saint Lucia; Web: www.stress-and-behavior.com

FEBRUARY 2017 SLAS 2017 - Society of Laboratory Automation and Screening. Feb 4-8; Washington, DC, USA; Web: www.slas.org Labquality Days2017. Feb 9-10; Helsinki, Finland; Web: www.labquality.fi

MARCH 2017 Pittcon Conference and Expo 2017. Mar 59; Atlanta, GA, USA; Web: http://pittcon.org KIMES 2017. Mar 16-19; Seoul, Korea; Web: www.kimes.kr ARABLAB 2017. Mar 20-23; Dubai, UAE; Web: www.arablab.com MEDLAB Asia Pacific 2017. Mar 29-31; Singapore; Web: www.medlabasia.com

APRIL 2017 ENDO 2017 – Endocrine Society Annual Meeting. Apr 1-4; Orlando, FL, USA; Web: www.endocrine.org MEDICAL FAIR INDIA. Apr 6-8; New Delhi, India; Web: http://medicalfair-india.com ECCMID 2017. Apr 22-25; Vienna, Austria; Web: www.eccmid.org/eccmid_2017

MAY 2017 Biomarkers & Diagnostics World Congress 2017. May 2-4; Philadelphia, PA, USA; www.biomarkerworldcongress.com ISLH 2017 – International Society of Laboratory Hematology. May 4-6; Honolulu, HI, USA; Web: www.islh.org LABVOLUTION 2017. May 16-18; Hanover, Germany; Web: www.biotechnica.de Diagnostica / Hospitalar 2017. May 16-19; Sao Paulo, Brazil; Web: www.hospitalar.com 19th European Congress of Endocrinolo-

gy. May 20-23; Munich, Germany; Web: www.ese-hormones.org/meetings ESPID 2017- European Society for Paediatric Infectious Diseases. May 23-27; Madrid, Spain; Web: www.kenes.com/ espid_2017_lp ESHG 2017 - European Human Genetics Conference. May 27-30; Copenhagen, Denmark, Web: www.eshg.org

JUNE 2017 EuroMedLab 2017. June 11-15; Athens, Greece; Web: www.ifcc.org FOCIS 2017 - Federation of Clinical Immunology Societies. June 14-17; Boston, MA, USA; Web: www.focisnet.org EAACI 2017 - European Academy of Allergy and Clinical Immunology . June 1721; Helsinki, Finland; Web: www.eaaci.org 2017 BIO International Convention. June 19-22; San Diego, CA, USA; Web: http:// convention.bio.org

JULY 2017 FEMS 2017- 7th Cong. of European Microbiologists. July 9-13; Valencia, Spain; Web: www.fems-microbiology2017.kenes.com AACC 2017 – 68th Annual Meeting of American Association for Clinical Chemistry. July 30-Aug 1; San Diego, CA, USA; www.aacc.org

AUGUST 2017 FIME 2017 - Florida International Medical Exhibition. Aug 1-3; Miami, FL, USA; Web: www.fimeshow.com

SEPTEMBER 2017 ESP 2017 - 29th European Congress of Pathology. Sep 2-6; Amsterdam, Netherlands; www.esp-congress.org Eurotox 2017 – 53rd Congress of the European Societies of Toxicology. Sep 1013; Bratislava, Slovakia; Web: www. eurotox2017.com 10th International Meeting of Pediatric Endocrinology. Sep 14-17; Paris, France; Web: http://internationalmeeting2017.org COLABLIOCLI 2017. Sep 17-20; Punta del Este, Uruguay; Web: http://colabiocli2017uy. com/punta_este.html 13th EFLM Symposium for Balkan Region. Sep 21-22; Belgrade, Serbia.

OCTOBER 2017 41st European Congress of Cytology. Oct 1-4; Graz, Austria; Web: www.efcs.eu 8th International Conference and Exhibition on Analytical & Bioanalytical Techniques. Oct 16-18; Milan, Italy; Web: http://analytical-bioanalytical.pharmaceuticalconferences.com ASHG 2017 - The American Society of Human Genetics. Oct 17-21; Orlando, FL, USA; Web: www.ashg.org IFCC WorldLab 2017. Oct 22-25; Durban, South Africa; www.ifcc.org

NOVEMBER 2017 MEDICA 2017. Nov 13-16; Dusseldorf, Germany; Web: www.medica.de AMP 2017 - Annual Meeting for Association for Molecular Pathology. Nov 16-18; Salt Lake City, UT; USA; Web: www.amp.org LabMedica International November/2016

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Vol. 33 No.7 • 11/ 2016 Advertiser

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107

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163

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173

Vircell . . . . . . . . . . . . . . . . . . . . .42

129

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