ARTICLE pubs.acs.org/JACS
Transduction of Glycan!Lectin Binding Using Near-Infrared Fluorescent Single-Walled Carbon Nanotubes for Glycan Profiling Nigel F. Reuel,† Jin-Ho Ahn,† Jong-Ho Kim,‡ Jingqing Zhang,† Ardemis A. Boghossian,† Lara K. Mahal,§ and Michael S. Strano*,† †
Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, United States Department of Chemical Engineering, Hanyang University, Ansan 426-791, Republic of Korea § Department of Chemistry, New York University, New York, New York 10003, United States ‡
bS Supporting Information ABSTRACT: There is significant interest in developing new detection platforms for characterizing glycosylated proteins, despite the lack of easily synthesized model glycans or high affinity receptors for this analytical problem. In this work, we demonstrate a sensor array employing recombinant lectins as glycan recognition sites tethered via Histidine tags to Ni2+ complexes that act as fluorescent quenchers for semiconducting single-walled carbon nanotubes (SWNTs) embedded in a chitosan hydrogel spot to measure binding kinetics of model glycans. We examine, as model glycans, both free and streptavidin-tethered biotinylated monosaccharides. Two higher-affined glycan!lectin pairs are explored: fucose (Fuc) to PA-IIL and N-acetylglucosamine (GlcNAc) to GafD. The dissociation constants (KD) for these pairs as free glycans (106 and 19 μM, respectively) and streptavidin-tethered (142 and 50 μM respectively) were found. The absolute detection limit for the current platform was found to be 2 μg of glycosylated protein or 100 ng of free glycan to 20 μg of lectin. Glycan detection (GlcNAc-streptavidin at 10 μM) is demonstrated at the single nanotube level as well by monitoring the fluorescence from individual SWNT sensors tethered to GafD lectin. Over a population of 1000 nanotubes, 289 of the SWNT sensors had signals strong enough to yield kinetic information (KD of 250 ( 10 μM). We are also able to identify the locations of “strong transducers” on the basis of dissociation constant (four sensors with KD < 10 μM) or overall signal modulation (eight sensors with >5% quench response). We report the key finding that the brightest SWNTs are not the best transducers of glycan binding. SWNTs ranging in intensity between 50 and 75% of the maximum show the greatest response. The ability to pinpoint strong-binding, single sensors is promising to build a nanoarray of glycan!lectin transducers as a high throughput method to profile glycans without protein labeling or glycan liberation pretreatment steps.
1. INTRODUCTION Understanding the interactions between glycans and carbohydrate recognition domains (CRDs) found on cellular and protein surfaces is vital to the fields of glycobiology, immunology, and pharmacology. Glycans decorating the surface of proteins substantially influence function, such as folding pathways, signaling, retention, and pharmacokinetics.1,2 The efficacy of protein-based therapeutics is largely dictated by their glycosylation,3 and thus de novo design of drugs that interact with known CRD sites, such as cell-adhesion modulating galectins,4 requires a greater understanding of the kinetic parameters between glycans and CRDs. Despite this importance, robust assays of protein glycosylation are underdeveloped, with the dominant profiling technologies falling to frontal affinity chromatography5,6 and mass spectrometry.7!9 Recently, the concept of the lectin microarray has emerged as a promising approach to investigating glycan-lectin interactions. These arrays take advantage of multivalent interactions to overcome the weak monovalent binding of lectin-glycan pairs (typically 10!7 M < Kd < 10!3 M) but are limited in their ability to transduce weak r 2011 American Chemical Society
mono- or multivalent interactions and also require fluorescence labeling.5,10!14 An emerging concept5,10 is to use multivariate responses of glycan binding to a library of lectins to discern their identity, but this requires detection methods that are necessarily sensitive enough to transduce the presence of weakly bound proteins. Label-free methods, which reduce sample volume requirements, have a distinct advantage in this approach by decreasing the absolute detection limit. Herein, we develop a fluorescent single-walled carbon nanotube (SWNT) sensor15 for glycan!lectin interactions with the ultimate goal of profiling glycans. Our approach is to couple band gap fluorescent SWNTs to receptor lectins, which are a host of naturally occurring carbohydrate binding proteins.16 We benchmark the sensor by evaluating the kinetic parameters between the anti-His tag antibody and comparing it to literature surface plasmon resonance (SPR) parameters. We then demonstrate the detection of fucose (Fuc) Received: August 16, 2011 Published: October 04, 2011 17923
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Journal of the American Chemical Society to PA-IIL lectin and N-acetylglucosamine (GlcNAc) to GafD lectin. Kinetic parameters are obtained by first measuring the fluorescence intensity of a large spot of SWNT, and then it is shown how the same signal can be increased by probing individual SWNT sensors and determining which sensors are most responsive to glycosylated analyte addition. Our objective of glycan profiling makes this effort distinct from recent work creating glycosolate carbon nanotubes for therapeutic purposes17,18 and electronic FET sensors for lectin, but not glycan, binding.19!21 In contrast, the aim of our work is to profile glycans via their selective binding to a fluorescent nanotube surface, a concept not yet demonstrated in the literature to date. Kinetic information on glycans!CRD interactions is currently determined by two types of analytical methodology: equilibrium and nonequilibrium.22 Nonequilibrium methods yield relative binding information rather than physical kinetic rates; that is, they specify which glycan!CRD combinations bind with greater or lesser affinity in reference to each other. These methods include ELISA,23 glycan microarrays,24 agglutination,25 and electrophoresis.26 A notable exception in this category of relative binding assays are some of the carbohydrate arrays from the CH Wong group that can yield quantitative fluorescent binding curves similar to SPR (below) and be translated to KD values.14 Equilibrium dialysis can determine the forward reaction rate (kf) of glycan!CRD binding but requires a large amount of glycan reagent, which is often a significant investment of time and money if complex sugars are used. Equilibrium titration calorimetry27,28 is a delicate technique to determine kinetic parameters from thermodynamic information but is rarely employed because of time and reagent expenses. Another equilibrium technique, frontal affinity chromatography,29 can be used to determine the affinity constant (KD) for most glycan!CRD pairs (KD > 10!7); however the glycan must be labeled for detection. The current standard for obtaining kinetic information from label-free groups is surface plasmon resonance (SPR) machines, such as the Biacore systems. In the case of glycan!CRD interactions, SPR can detect both the forward and reverse kinetic rates for a wide range of affinities (KD: mM!pM range). However, to induce a detectable signal the analyte must have significant mass. Thus glycans are typically immobilized on the gold surface (often using neoglycoproteins,30 i.e. glycans synthetically coupled to a protein backbone), and the more substantial lectins are used as the binding analytes. This can bias the analysis of single lectin! glycan interactions as presentation and density of the glycan is a critical parameter in CRD binding, and the immobilization methodology can alter this.31 Though more difficult, lectins can also be immobilized on the SPR surface and detect glycosylated analytes, but again these must have enough mass to transduce a change in refractive index. We show that the SWNT-based fluorescence sensors developed in this work demonstrate loading curve signals competitive with SPR, in both shape and analysis technique, but they differ in a few significant ways. First, the detection scheme is reversed. The lectins are the tethered sensors, and the glycans are the analyte in solution. This allows us to determine the kinetics of free glycans as well as glycoproteins giving us precise control over carbohydrate presentation in the interaction. Second, the amount of analyte needed for each experiment (2 μg of glycosylated protein or 100 ng of free glycan) is smaller than what is necessary for SPR experiments, which require analyte flow to overcome mass-transport effects (at its best, SPR requires 20!250 μg of analyte protein32). Third, each SWNT-sensor spot
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can be tethered to different lectins and illuminated simultaneously, creating the potential for a multiplexed, quantitative detection of analyte binding analogous to an array-reading SPR machine with distinct advantages in sample size and run time.
2. EXPERIMENTAL SECTION 2.1. Lectin Expression. The resulting plasmid pET41GafD and pET41-PAIIL were transformed into BL21(DE3) Star (Invitrogen) according to standard procedures.33 A single colony was used to inoculate 5 mL of LB medium containing kanamycin (50 μg mL!1). 3 mL of the overnight culture at 37 !C were used as an inoculum to a 350 mL flask of LB containing 50 μg mL!1, and this was incubated again with shaking at 37 !C. Heterologous protein was induced by the addition of isopropyl β-D-1-thiogalactopyranoside (IPTG, final concentration 1 mM) once this culture had reached log phase (A600 of 0.6). Growth was continued for 6 h before the cells were harvested by centrifugation. For His-tag protein purification, a harvested cell pellet was washed twice with phosphate-buffered saline, PBS (10 mM, pH 7.4), and then lysed with Complete Lysis-B (Roche Applied Science). The crude lysate was clarified by centrifugation prior to application to a 3 mL Ni-NTA agarose column (Qiagen). Nonspecifically bound proteins were removed from the column with wash buffer (50 mM NaH2PO4, pH 8.0, 300 mM NaCl, 20 mM imidazole), and bound His-tag GafD and PAIIL were eluted with elution buffer (50 mM NaH2PO4, pH 8.0, 300 mM NaCl and 250 mM imidazole). Eluted samples were analyzed by 15% SDS-PAGE, and the protein concentration was determined with a BCA assay kit according to manufacturer’s instructions (Pierce). To change the buffer with PBS (10 mM, pH 7.4), the eluted solution was centrifuged through a centrifugal filter with a molecular cutoff of 10 kDa (Millipore), and the concentration of lectin was finally adjusted to 4 mg mL!1. 2.2. Glycan and Model Glycoprotein Probes. Biotinylated glycans were provided by the Consortium of Functional Glycomics ! Scripps Institute Group. For this work, glycans B121 (GlcNAcβ-SpNHLC-LC-Biotin) and B158 were used (Fucα1-2Galβ1-4Glcβ-SpNH-LCLC-Biotin) where Sp is 2-azidoethyl and LC-LC biotin is a standard biotin reagent with an extra-long spacer group (Pierce ID# 21343). The lyophilized sugars were dissolved in 3 mL of 1" PBS to create stock solutions and stored at !20 !C. To construct model glycoprotein probes, the biotinylated glycans were incubated with streptavidin (Sigma Aldrich SO677) for 1 h at 20 !C in a 6:1 molar ratio to allow maximum binding to the four biotin binding sites on each streptavidin. Excess biotinylated glycans were filtered away from the glycoproteins by centrifugation through an Amicon filter (16 300g on Labnet Inc. centrifuge, 10 min, 30 000 kDa cutoff, Milipore). The glycoproteins were washed on the filter 3" (400 μL PBS) and then resuspended in PBS at the desired concentrations. 2.3. Native PAGE Binding Analysis. The interaction between lectin and glycan was analyzed by 15% native PAGE according to the method of Sch€agger and von Jagow with minor modifications.34 Briefly, electrophoresis was performed using a Mini Protean Electrophoresis system (Bio-Rad, USA) under nondenaturing conditions to examine molecular interactions. Nonreduced protein/glycan samples (GafD Lectin held constant at 30 μg per lane, GlcNAc-Strept added at 5, 10, 20, and 30 μg) in the sample buffer (20% glycerol, 200 mM Tris-HCl, pH 6.8, 0.05% bromophenol blue) were applied to the gel (gel buffer: 25 mM Tris-HCl, 200 mM glycine). Electrophoresis was performed at 80 V for 120 min. After electrophoresis, the protein bands were visualized by staining with Coomassie Brilliant Blue R-250. 2.4. Construction of Chitosan-SWNT Sensor Chips. To increase the reproducibility of our sensors, we have introduced a careful automated printing method of the chitosan gel (Supporting Figure 1). Patterned glass microscope slides (Tekdon) were inserted in a microarray printer (Digilab MicroSys System) where the robotic head was 17924
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Scheme 1. Processing Steps To Tether a His Tag Sensor Protein to SWNTa
a
(1) Wrap SWNT in chitosan, (2) functionalize chitosan with carboxylic acids, (3) attach tethered NTA group, (4) chelate nickel with NTA, (5) add His tag sensor protein to SWNT.
programmed to dispense alternating layers of chitosan-SWNT (0.25 wt % chitosan (CHI), 1 vol% acetic acid, 30 μg/mL suspended (6,5) SWNT) and cross-linker (10 vol% glutaraldehyde). The suspended SWNT was made from Southwest Nanotechnologies, Inc. CoMoCAT nanotubes sonicated in 0.25 wt % chitosan (CHI) and 1 vol% acetic acid for 45 min at 40% amplitude with a probe!tip sonicator (Cole Parmer, Model CV18). For each sensor spot 10 alternating layers of SWNT-CHI and cross-linker were printed at 100 nL per layer, resulting in a highly uniform gel of 1 μL of SWNT-CHI material. The chips were printed in a humidified enclosure (85% RH) at 25 !C and allowed to cross-link overnight in the same environment. Nickel-NTA groups were introduced as previously reported.15 Briefly, the chips were washed with a dilute basic buffer (0.01 M NaOH) and water three times. Carboxylic acid groups were introduced to the chitosan wrapped SWNT by bathing the chips in succinic anhydride (0.1 M) overnight (Scheme 1, step 2). The chips were then washed three times with water, and the carboxylic acid groups were activated via N-(3- dimethylaminopropyl)-N0 -ethylcarbodiimide hydrochloride (EDC, 0.1 M) and N-hydroxysuccinimide (NHS, 0.1 M). The chips were bathed in this solution for 2 h at 25 !C and examined for the expected formation of bubbles. The chips were again washed and allowed to bathe in a solution of a linked tricarboxylic acid group (Nα,Nα-bis(carboxymethyl)-L-lysine aka NTA, 33 mM) overnight (Scheme 1, step 3). The chips were washed in water again and stored in a 100 mM solution of NiSO4 to allow maximum binding of nickel to the NTA chelating groups (Scheme 1, step 4). Thus in each experiment the SWNT sensors start in their maximally quenched state due to the close proximity of the nickel. During experimentation the excess NiSO4 is washed away with water and the sensor protein, Histagged lectin, is tethered to the SWNT sensors via the chelated nickel group (Scheme 1, step 5). The large protein groups cause the nickel group to move away from the SWNT sensor, due to steric loading (discussed below), and part of the quenched fluorescent signal returns. 2.5. Ensemble Measurements of Sensors. A custom-made near-infrared inverted microscope (Zeiss D.1 Observer) setup allows us to probe the fluorescent emissions of our SWNT sensors (Figure 1a). The chips are secured on the microscope stage, and the objective (50"/ 0.7 Zeiss) is pushed into contact with a blank portion of the glass slide (no SWNT-CHI) to obtain a 5 s background spectrum, which is subsequently subtracted from the response spectra. The objective is then moved under an SWNT-CHI gel spot and again pushed in contact with the glass slide. By placing the objective in the maximum z-axis position, the microscope can image a higher plane of the SWNT-CHI gel
where more analyte response is observed. The SWNTs are excited by a 785 nm laser (B&W Tek, 495 mW), and the emission is sent to a spectrometer (Princeton Instruments Acton SpectraPro 2500i Spectrograph) and accompanying nIR camera (Intervac MOSIR Camera 350). The spectra are collected via WinSpec software (Princeton Instruments) and analyzed with custom Matlab (Mathworks) code. To maximize signal stability, the spectrometer is cooled with liquid nitrogen 2 h prior to experimentation and the laser is allowed to reach peak stability for 2 h. The SWNT-CHI gel has a small transient region when first exposed to the laser due to local heating and further permeation of Ni2+ in the gel; thus each spot is pre-exposed to the laser for 5 min before data are gathered. Data are gathered in the form of emission intensity spectra (950!1250 nm) integrated for 5 s. A typical experiment was run for approximately 1000 frames (at 5 s each) and included a few addition and washing steps to detect lectin! glycan binding (Figure 1b). First NiSO4 was again added to ensure that the SWNT sensors were responsive and that the NTA chelating groups were fully loaded with Ni2+ groups. The nickel was then washed away with PBS three times, leaving ∼20 μL of PBS on the sensors. The His-tag lectin was then added to the sensor (20 μL at 2 mg/mL) and allowed to bind for 300 s. Excess lectin was again washed by PBS three times, and 20 ( 1 μL PBS were left to bathe the sensor. The sensor was allowed to equilibrate for 100 s, and 20 μL of analyte were added. The analytes tested in this paper include free biotinylated glycans, glycans tethered to streptavidin, and anti His-tag antibody. Each of these, upon binding, causes an increase in SWNT luminescence (Figure 1b). The sensor response was recorded for 500 s. It is essential to record stabilization frames before and after the analyte addition in order to correct for any focal drift caused by the tension of the objective in contact with the glass slide (see Results section). 2.6. Single SWNT Sensor Measurements. A second custom microscope was used to collect emission intensities of single SWNT sensors. The SWNT-CHI gel was diluted to 3 μg/mL of SWNT and spin-coated (3000 rpm for 30 s on Laurell Technologies Corporation, WS-650MZ-23NPP/LITE) on glass-bottom Petri dishes (MarTec Corp). The Petri dishes were then placed on the microscope platform (Zeiss D.1 Observer), and the oil-immersed objective (Zeiss 100"/1.46) was focused on the SWNT sensors on the glass surface. The SWNTs were excited by a 660 nm laser (Crystal Laser, 100 mW), and the emission intensities were recorded by a nitrogen-cooled InGaAs array (Princeton Instruments). Again WinSpec software (Princeton Intrsuments) was used to collect the data in the form of a stacked Tiff 17925
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International Scholarly Research Network ISRN Nanotechnology Volume 2012, Article ID 102783, 9 pages doi:10.5402/2012/102783
Review Article A CNTFET-Based Nanowired Induction Two-Way Transducers Rostyslav Sklyar Verchratskogo st. 15-1, Lviv 79010, Ukraine Correspondence should be addressed to Rostyslav Sklyar, sklyar@tsp.lviv.ua Received 15 December 2011; Accepted 28 February 2012 Academic Editors: C. A. Charitidis and J. Sha Copyright Š 2012 Rostyslav Sklyar. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. A complex of the induction magnetic field two-way nanotransducers of the different physical values for both the external and implantable interfaces in a wide range of arrays are summarized. Implementation of the nanowires allows reliable transducing of the biosignals’ partials and bringing of carbon nanotubes into circuits leading to examination of the superconducting transition. Novel sensors are based on the induction magnetic field principle, which causes their interaction with an ambient EM field. Mathematical description of both the signal and mediums defines space embracing of the relevant interfacing devices. As a result, a wide range of the nano-bio-transducers allow both delivering the variety of ionized biosignals and interface the bioEM signals with further stages of electronic systems. The space coverage and transducing values properties of the state-of-the-art magnetic interfaces are summarized, and directions for their future development are deduced.
1. Introduction: Biophysical Signals, Transducing, and Interface Applications A biosensor is a device that incorporates a biologically active layer as the recognition element and converts the physical parameters of the biological interaction into a measurable analytical signal [1]. Understanding of biosignals’ (BS) nature and properties of their mediums are a basis for effective design of magnetic interfaces (MIs). Rapid progress in the advancement of several key science areas including nanoscale interfaces has stimulated the development of electronic sensor technologies applicable to many diverse areas of human activity. For example, the conceptualization and production of electronic nose devices have resulted in the creation of a remarkable new sector of sensor technology resulting from the invention of numerous new types of olfactory-competent electronic sensors and sensor arrays [2]. The growing variety of biosensors can be grouped into two categories: implantable and external. In turn, the last one has two existing paradigms: wearable sensor and noncontact sensor. A wearable sensor had potential to be intrusive, and noncontact sensor methods may still be intrusiveness to a certain extent, while a noncontact sensor is limited in its capability of acquiring physiological signals [3]. Voltage potentials of the living organism and its organs are measured
by both implantable and external electric field probes of high sensitivity [4]. Information on organ activity is obtained by measuring biomagnetic signals. For such purposes a multichannel high-temperature superconducting quantum interference device (high Tc SQUID) system for magnetocardiography (MCG) and magnetoencephalography (MEG) of humans, with high magnetic field resolution, has been developed [5, 6]. The most current sensing devices give us the possibility to receive a full scale of both the internal and external control BS. The internal ones are picking up by polymeric microprobes, CMOS chips, and nanoneedles, while the external by electromyography and neuroprosthetic (electroencephalogram (EEG) and MEG) systems. Improving an informational capability of the interface is implemented by the application of the advanced superconducting transducer and electromagnetic (EM) transistor/memristor [7, 8]. These elements are arranged into the arrays of a different configurations and can cover the order of spaces from macro- to nanolevels. There are a number of methods and devices for transducing different BS into recordable or measurable information. The transfer of nerve impulses (NI) is the main data flow that carries sensory information to the brain and control signals from it and from the spinal cord to the limbs.
2 Moreover, the complex view on BS requires further stages of precise processing in order to decode the received or control information. There are different kinds of transducers/sensors for picking up NI: room-temperature and superconducting, external, and implantable. Development of such devices is increasing the penetration into bioprocess while simultaneously simplifying the exploitation of the measuring systems in order to bring them closer to the wide range of applications. For this reason, the magnetometer with a room-temperature pickup coil (PC) for detecting signals, which can clearly be detected in higher frequency range, was developed in order to simplify the SQUID system. The PC is set outside the cryostat and is connected to the input coil of the SQUID [9] or a channel of superconducting field-effect transistor (SuFET) [10]. On the other hand, implantableinto-nerve fiber transducers are evolving from the ordinary Si-chip microelectronics devices [11] into superconducting and nanodevices [12, 13]. The recent achievements in nanoelectronics can be regarded as a further step in the progress of BS transduction. They give us the possibility to create the most advanced and universal device on the basis of known microsystems. Such a sensor/transducer is suitable for picking up BS— NI, electrically active (ionized) molecules, and the basepair recognition event in DNA sequences—and transforming it into recognizable information in the form of electric voltage, or a concentration of organic or chemical substances. Moreover, this process can be executed in reverse. Substances and/or voltages influence BSs, thereby controling or creating them (BS) [14]. Steady and rapid progress in the robotics field requires ever quicker and better human-machine interaction and the development of a new generation of interfaces for intelligent systems. Such advances give rise to markedly increased biophysical research on the one hand and the need for new bioelectronic devices on the other. Transduction and measurement of BS are key elements of MIs design. There are two means involved in signal transduction: (1) biochemical—by hormones and enzymes; (2) biophysical— by nerve impulses (ionic currents). Let us consider the biophysical ones as useful for the said interfaces design above. There are two values—voltage and electric current—which characterize the pathway of transduction [15]. Calculations of PC arrays were performed with the primary sensor flux transformer sites distributed uniformly on a spherical sensor shell, extending from the vertex to a maximum angle max [16]. The radial magnetometers and gradiometers occupy one site each, there are two orthogonal planar gradiometers at each site and there are three orthogonal magnetometers at each site for vector magnetometers. Coverage can be achieved by designing some kind of density control mechanism, that is, scheduling the sensors to work alternatively to minimize the power wastage due to the overlap of active nodes’ sensing areas. The sensing area of a node is a disk of a given radius (sensing range). The sensing energy consumption is proportional to the area of sensing disks or the power consumption per unit [17]. There are two broad ways of brain-computer interface (BCI): invasive and noninvasive. The invasive technique can capture intracortical action potentials of neurons and thus,
ISRN Nanotechnology provides high signal strength spatiotemporally, for example, prediction of movement trajectory. In noninvasive technique, EEG and MEG have emerged as viable options; both of them have time resolutions in milliseconds. Any activity in brain is accompanied by change in ionic concentrations in neuron leading to polarization and depolarization. Such an electrical activity is measured by EEG, while MEG measures the magnetic field associated with these currents. Electric and magnetic fields are oriented perpendicular to each other [18]. Application of organic-, chemical-, and carbonnanotubes- (CNT-) based FETs for design of the superconducting transducers (SuFETTrs) of BS into different quantities (electrical and biochemical) is the proposed variant of interfacing [19]. The placement of the devices can be carried out both in vivo and in vitro with the possibility of forming the controlling BS from the said quantities. The range of picked up BS varies from 0.6 nA to 10 µA with frequencies from 20 to 2000 Hz. A further step should be the synthesis of the said two methods in order to develop the internal (implantable) nano-bio-interface arrays. This means wrapping of molecular nanowired PC around the axons of a nerve fibre or synapses of neurons in order to obtain the natural biosignals from the nervous system and brain. This leads to sensing access across a vast range of spatial and temporal scales, including the ability to read neural signals from a select subset of single neural cells in vivo. Moreover, this process can be executed in reverse for introducing the artificial control signals with the local neural code into the single cell electrical activity.
2. Biosignals and Nanoelements for Their Transduction As an electrical signal, the biosignal has two components: electrical potential or voltage and ionic or electronic currents. The first component is sufficiently developed and does not require penetration into the substances of biosignal propagation. The marketable progress in transducing of the second component began when the necessary instrumentation for measurement of micro- and nanodimensions had been created [14]. Short platinum nanowires (NWs) already have been used in submicroscopic sensors and other applications. A method of making long (cm) Pt NW of a few nanometers in diameter from electrospinning was described [20]. Those wires could be woven into the first self-supporting webs of pure platinum. Double-gated silicon NW structures (DGSiNW), where the position and/or type of the charge could be tuned within the NW by electric field, have been studied [21]. Self-assembled molecular nanowires were found to be composed of a single crystal, allowing good electrical transport with low resistivity [22]. An interesting structure is that of helical CNT or nanocoils for PCs. Nanocoils offer unique electronic properties that straight CNT do not have. The plasticity of CNT will be relevant to their use in nanoscale devices [23]. Carbon nanocoils (CNCs), as a new type of promising
ISRN Nanotechnology nanomaterials, have attracted considerable attention because of their potential applications, such as parts for nanoor micro-electromechanical systems, EM wave absorbers, reinforced composites, nanosolenoids, and field emission devices [24] Integrated CMOS image sensor device for in vivo neural imaging has been developed. Improvement in the packaging process has resulted in a compact single-chip device for minimally invasive imaging inside the mouse brain [25]. Application of the SuFET’s modifications such as CMOSuFET (low Tc ) [26] and coplanar SuFET (high Tc ) [27] broadens the range of requirements, which are being satisfied by the SuFETTr. Alternatively, an FET-based neurotransducer with CNT or PC kind of input circuit for the nerve and neuron impulse has been designed. A CNTFET with a high-temperature superconducting channel is introduced into the nerve fibre or brain tissue for transducing their signals in both directions [28]. Flexible antennas have the potential to enhance the emerging field of flexible electronics, which is primarily motivated by the desire to incorporate electronics into flexible substrates such as textiles, displays, and bandages [29]. The ability to reversibly deform antennas may also enable new capabilities (e.g., rolling and unrolling for remote deployment, enhanced durability). Relative to conventional copper antennas, fluidic antennas have several advantages [30]. Furthermore, it has been shown that ultrathin layers of metal can display superconductivity, but any limits on the size of superconducting systems remain a mystery. On the other hand, (BETS) 2GaCl4, where BETS is bis (ethylenedithio) tetraselenafulvalene, is an organic superconductor, and in bulk it displays a superconducting gap that increases exponentially with the length of the molecular chain [31]. Graphene-solution-gated FET (G-SGFET) fabricated on copper foil offers outstanding electronic performance, is chemically stable and biologically inert, and can readily be processed on flexible substrates. Not only were the “action potentials” of individual cells detectable above the intrinsic electrical noise of the transistors, but these cellular signals could be recorded with high spatial and temporal resolution. The analysis of the recorded cell signals and the electronic noise of the transistors confirm that graphene transistors surpass state-of-the-art devices for bioelectronic applications [32]. An organic FET (OFET) is characterized by textile process fully compatible size and geometry. This transistor has shown very interesting performances, with typical values of the electronic parameters very similar to those of planar devices. This result is very promising in view of innovative applications in the field of smart textiles [33]. Also FETs with single- and doublewire channels (NWFET) were fabricated to give some indication of the potential application of the molecular wires [22]. Finally, inkjet-printed FETs using carboxyl-functionalized nanotubes as source, drain, and gate electrodes, poly (ethylene glycol) (PEG-) functionalized nanotubes as the channel, and PEG as the gate dielectric were also tested and characterized. Considerable nonlinear transport in conjunction with a high channel current on/off
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Figure 1: Diagnostics of the biomedium with the necessary drugs delivering.
ratio of 70 was observed with PEG-functionalized nanotubes. The positive temperature coefficient of channel resistance shows the nonmetallic behavior of the inkjet-printed films [34]. Finally, FETs with single- and doublewire channels were fabricated to give some indication of the potential application of the molecular wires. Substantial progress has been made in defining the performance limits and exploring applications based on NWFETs [22]. A five-channel FET structure is composed of two double-gate transistors and a bottom single-gate transistor on a silicon-on-insulator. 3D transistor structures such as multiple-gate FETs have been proposed and extensively studied as a promising solution to overcome the scaling limitations of planar bulk devices. They offer excellent multigate control of the channels and higher current drive [35]. In high-performance n-channel OFETs, charge carrier injection at the interface between the organic film and source/drain electrodes plays a crucial role [36].
3. An SuFETTr-Based Devices
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The advent of semiconductor devices with nanoscale dimensions creates the potential to integrate nanoelectronics and optoelectronic devices with a great variety of biological systems. In such a case, it is possible to substitute the microcomputer in an object-oriented problem solution scheme by the natural processing organ-brain or spinal cord. As a result, the software component will be eliminated and the most general characterization of the measurement problem in one-coordinate-dimensional measurements could be acquired naturally, according to the feedback reaction on the input exposure. Moreover, the organs of the senses of living beings could be regarded in the same way as the natural
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Figure 2: Transducing of the nerve impulses and introducing of the relevant artificial signals [15].
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Figure 3: PC for “two-dimensional” gradiometer that detects both axial-second-order gradient and planar-first-order gradient of MF [37]. Copyright 2007 The Japan Society of Applied Physics.
biosensors of the relevant physical and chemical quantities [19, 39]. The proposal to measure the biosignal values of different origins with advanced nanosensors of EM quantities is justified when allowing for superconducting abilities of the devices. They are composed in full-scale arrays. The said arrays can be both implantable into ionic channels of an organism and sheathed on the sources of the EM emanation. Nanowired head sensors function both in passive mode for picking up the biosignals and with additional excitation of a defined biomedium through the same head (in reverse) [40– 42]. 3.1. The Arrangements of a NanoFET-Based Delivery and Transducing. The advances in nanotechnology are opening
Figure 4: Comparison of the discrete geometry of three selfassembly models. (a) Unfolding an HP chain. (b)–(d) Several representations of unfolding a cube [38]. Copyright 2011 National Academy of Sciences, USA.
the way to achieving direct electrical contact of nanoelectronic structures with electrically and electrochemically active subcellular structures, including ion channels, receptors, and transmembrane proteins. The method of combining the bioelectric nature of the nerve impulses NI and synaptic currents between neighboring neurons with body-temperature PC and zero-resistance-CNT-based input of the SuFET device in order to obtain most advantageous biosensor/transducer was recently advanced [43]. On the other hand, neuroelectronic systems for two-way interfacing of the neuronal and the electronic components by capacitive contacts and by FETs with an open gate were developed. A nanoSuFET with a high-temperature superconducting channel is introduced into the nerve fibre or brain tissue for transducing their signals in both directions. Such a sensor/actuator is suitable for picking up BS—NI and electrically active (ionized) molecules—and transforming it into recognizable information in the form of electric
ISRN Nanotechnology
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Figure 5: Interaction of the natural and artificial processing bodies trough a SuFETTr-based interface.
voltage, or a concentration of organic or chemical substances (Figure 1). Moreover, this process can be executed in reverse. Substances and/or voltages influence BS, thereby controlling or creating the said media (Figure 2). As a result, we have achieved SuFETTr that is suitable for ascertaining the variety of values. Two directions of SuFETTr function enable decoding of the NI by comparing the result of its reaction on some process or organ with an action on them of the simulated electrical or biochemical signal after their reverse transducing through the SuFETTr(s) [44]. 3.2. Arrangement of PSs in Arrays. An MF sensing array consists of primary sensors and, optionally, reference sensors. Primary sensors use flux transformers located in a close proximity to the scalp or chest surface, where they couple to the brain’s or heart’s MFs, respectively [7, 45]. The reference sensors are used to subtract the environmental noise from the primary sensor outputs. The flux transformer design dictates its relative sensitivity to near and distant sources. Thus, the primary flux transformers can, in addition to detecting the brain signals, also provide various degrees of the environmental noise rejection. Flux transformers (magnetometers) have the highest sensitivity to both near- and far-field sources. Thus, they do not reduce the environmental noise (and must rely solely on the references or other techniques for the noise cancellation) [16]. Wire-wound gradiometers are the most conventional and are commercially available for EM systems. The PCs of wire-wound gradiometers are wound in opposition and balanced so that a uniform field links the zero net flux [37]. Conventional gradiometers, such as wire-wound, thin-film, or electronic gradiometers, are axial or planar, that is, “one dimensional” that detect the gradient of a MF in one direction. These one-dimensional gradiometers effectively reduce the ambient MF as their order increases. However, they also reduce the biomagnetic signal. A “twodimensional” gradiometer detects the gradient of an MF in
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Figure 6: Interfacing ability (power) of the specific input elements. It depends on two groups of space characteristics (form and direction) for the available transducing media.
two orthogonal directions to achieve high SNR. It focuses on a two-dimensional gradiometer that detects both the axialsecond-order and planar-first-order gradients of an MF. Figure 3 shows a PC for the two-dimensional gradiometer. The discrete configuration space is a graph. Each node in the graph corresponds to an intermediate, and its neighbors are intermediates to which it can be folded or unfolded [38]. A common feature shared by folding of hydrophobicpolar chains on a lattice and self-folding a net through vertex connections is that in both cases the process of folding is driven by the formation of secondary links between topological neighbors (Figure 4). The best 2D arrangements, called planar nets, to create self-folding polyhedra with dimensions of a few hundred microns are determined, and optimal configurations for creating 3D geometric shapes have been found. The importance of being able to address nanoscale elements in arrays goes beyond the area of nanocomputing and will be critical to the realization of other integrated nanosystems such as chemical/biological sensors. A regular crossed-NW FET array that consists of n-input Iin and moutput Uout NWs, in which outputs are the active channels of FETs and the inputs function as gate electrodes that turn these output lines on and off [46, 47]. 3.3. Multisensor Data Fusion from Arrays. A further step should be synthesis of the said two methods in order to develop the external (nonimplantable) MCG&MEG signalsto-processor connection. The EM sensors are surface PCs, which are used in regular configuration where PCs with a small distance between each other are distributed under the heart or brain surface to pick up the local signals within the place of interest. The problem of sensing the EM signal for amplification/switching/memory with a speed of light in a