SFM brochure

TM

The Journey to Discovery starts here. The Commitment to Excellence starts now.

Akron Biotech is a research-based biotechnology

company that manufactures and sells high quality

recombinant proteins, animal sera and serumfree media to cell biology and immunology researchers

in

academic,

corporate laboratories.

government,

and

Our world-class scientific advisory board and collaborations with leading research institutions helps Akron Biotech maintain the highest level of

technical expertise in researching, developing, and manufacturing the products that you depend on for your research.

As a truly global supplier, Akron Biotech exports products worldwide and is committed to providing

fast delivery and best-in-class technical support.

We understand the economic pressures on today’s busy laboratories, which is why we are one of a select few suppliers that provide bulk quantities of our products. Our staff recognizes the

need to deliver products of the highest quality on time and on budget.

Our “best practice” approach to manufacturing

guarantees batch-to-batch consistency for every product we sell. Akron Biotech identifies and

sources the highest quality raw materials. Our quality control department, which works closely with production, maintains thorough records of our entire supply chain, from source materials through finished product. In addition, our quality

assurance staff oversees customer service, technical support, production, and quality control

operations to assure that each customer’s needs are met.

Akron Biotech is headquartered in South Florida, with offices and manufacturing facilities around the world.

SERUM-FREE MEDIA

For Mammalian Cell Culture in Suspension

AK-Media hi, AK-Media mid, AK-Media lo, AK-Grow hi & AK-Grow lo

These formulations have been successfully used in: • Culture of myeloma and hybridoma cells • Monoclonal antibody production • Culture of human lymphocytes (including stimulated or transformed cells) AK-Media hi and AK-Media mid formulations contain no exotic growth factors and are therefore cost-efficient. The relatively higher protein content of AK-Media hi is aimed at maximizing cell growth, while the lower protein content in AK-Media mid represents a compromise between promoting cell growth and monoclonal antibody production, while easing purification.

AK-Media lo medium is totally free of bovine serum albumin.

Despite its protein content of <18 µg/ml, AK-Media lo is remarkably effective for growing a wide variety of hybridomas and lymphocytes.

AK-Media series are ready-to-use media, requiring only the addition of L-Glutamine and antibiotics.

AK-Grow hi and AK-Grow lo are serum-free supplements intended for customers who prefer to formulate their own final medium from the basal medium of their choice.

PRODUCT

AK-Media hi

SFM-High Level Protein w/o L-Glutamine

AK-Grow products are 50-fold concentrates and therefore are recommended for use at a concentration of 2% (although in some cases 1% maybe sufficient). DMEM: F-12 (1:1) has found to be most generally effective as the basal media, but many cell lines grow well with RPMI, DMEM or lscove’s.

AK-Media hi

10X Concentrate w/o L-Glutamine, w/o Sodium Bicarbonate AK-Media mid

SFM -Mid-Level Protein w/o L-Glutamine

Stability

AK-Media mid

10X Concentrate w/o L-Glutamine,

The AK-Media hi, mid and lo are stable for 12 months when stored at 2-8°C.

w/o Sodium Bicarbonate AK-Media lo

Low Protein Media BSA-Free,

The AK-Grow supplements are stable for 18 months when stored at -20°C.

w/o L-Glutamine AK-Media lo

10X Concentrate w/o L-Glutamine,

Adaptation of Cells

w/o Sodium Bicarbonate AK-Grow hi

For many cell types no adaptation procedures are necessary and may even be detrimental. In other cases standard weaning procedures may be necessary.

SFM Supplement - High Level Protein, 50X concentrate

For exact adaptation procedures please refer to our Method of Use page (available upon request).

AK-Grow lo

SFM Supplement - Low Level Protein,

To adapt the SFM to your specific conditions we suggest that you test samples of our various SFM products.

50X concentrate

03

Catalogue # SFM-HLP100

Size 100ml

SFM-HLP500

500ml

SFM-HLT100

100ml

SFM-HLT500 SFM-MLP100

500ml 100ml

SFM-MLP500

500ml

SFM-MLT100

100ml

SFM-LLP100

100ml

SFM-LLT100

100ml

SFM-MLT500

SFM-LLP500

SFM-LLT500 SFM-SPH002

500ml

500ml

500ml 2ml

SFM-SPH010

10ml

SFM-SPH100

100ml

SFM-SPH020

SFM-SPL002

SFM-SPL010 SFM-SPL020

20ml

2ml

10ml 20ml

SERUM-FREE MEDIA

For Adherent and Suspension Cultures

AK-MultiCell, AK-CHO adh, AK-CHO sus and AK-CHO supp AK-MultiCell is a ready-to-use formulation for adherent cells that contains no albumin and requires only the addition of L-Glutamine.

The protein content of AK-MultiCell is therefore <30mg/L, and the medium contains no growth factors or hormones other than insulin. The formulation also contains no attachment factor, which in many (but not all) cases must be added for successful use.

Stability

AK-MultiCell is stable for 12 months when stored at 2-8째C. Testing for the need for fibronectin or other additives

Since AK-MultiCell is intended for use with adherent cells, in many cases it will be necessary to use fibronectin (AK-Fibronectin) to induce initial adhesion of the cells to the growth surface. Each cell line must be checked individually, as some require fibronectin while others do not. In some instances fibronectin will improve performance but not strictly be required for growth.

In the cases of a few unusual types of cells it may also be necessary to add to the AK-MultiCell medium a specific growth factor or hormone, for which the cells have a specific requirement.

Growth of various anchorage dependent cells in AK-MultiCell as compared with conventional serum-supplemented medium* DMEM + 10% FBS

Cell

3T3 A-549 B16-F10 BGM BHK-21 BS-C-1 CEF HELA HEp-2 MA-10** VERO

Seeding Density/cm

5x10(3) 1x10(4) 5x10(3) 1x10(4) 2.5x10(4) 1x10(4) 1.2x10(4) 5x10(3) 5x10(3) 2.5x10(4) 5x10(3)

(2)

Doubling time (hours)

24.0 26.4 30.0 19.2 14.4 24.0 28.8 48.0 57.0 18.0 16.5

Maximum density/cm

AK-MultiCell

(2)

3.3x10(5) 4.5x10(5) 5.0x10(5) 4.0x10(5) 4.5x10(5) 2.8x10(5) ----------6.5x10(5) 5.5x10(5) 2.7x10(5) 4.1x10(5)

Additives

Bombesin SBTI*** Fibronectin AK-Fibronectin --------------------AK-Fibronectin --------------------AK-Fibronectin AK-Fibronectin AK-Fibronectin AK-Fibronectin

Doubling Time (Hours)

25.2 33.0 30.0 30.5 12.0 28.0 36.3 36.0 30.0 16.5 18.0

Maximum Density/cm

(2)

3.0x10(5) 2.8x10(5) 5.5x10(5) 3.4x10(5) 9.0x10(5) 1.9x10(5) ----------6.0x10(5) 6.5x10(5) 3.8x10(5) 3.8x10(5)

* DMEM+10% FBS: 3T3, A-549, BHK-21, BS-C-1, VERO, RPMI-1640+10% FBS: B16-F10, BGM, HELA, HEp-2, M-199/F10 (1:2): CEF / ** Cells do not grow with FBS but with 15% horse serum in RPMI / *** Soybean trypsin inhibitor

04

AK-CHO adh for adherent cultures is the basic SFM-base formulation for CHO cells.

AK-CHO sus is our basic SFM-base formulation for CHO cells in suspension.

Both media contain amino acids, vitamins, salts, lipids and trace elements They are intended for the growth of CHOK1 and transfected CHO cells containing recombinant DNA related to the DHFR gene.

Preparation of both formulations requires the addition of AK-CHO supp. The SFM supplement contains proteins and other components, essential for the media Stability

AK-CHO sus and AK-CHO adh are stable for 12 months when stored at 2-8째C. AK-CHO supp is a 100 fold concentrate and is stable for 18 months when stored at -20째C. PRODUCT

Catalogue #

Size

Multi Purpose SFM for Adherent Cells

SFM-MPM100

100ml

SFM-MPT100

100ml

SFM-ADH100

100ml

SFM-ADT100

100ml

AK-MultiCell

w/o L-Glutamine AK-MultiCell

10X Concentrate, w/o L-Glutamine, w/o Sodium Bicarbonate

AK-CHO adh SFM Base

For use with anchorage dependent CHO cells, w/o L-Glutamine AK-CHO adh SFM Base

10X concentrate, w/o L-Glutamine,

w/o Sodium Bicarbonate

AK-CHO sus SFM Base

For use with CHO cells in suspension,

SFM-MPM500

SFM-MPT500

SFM-ADH500

SFM-ADT500 SFM-SUS100

500ml

500ml

500ml

500ml 100ml

w/o L-Glutamine

SFM-SUS500

500ml

10X Concentrate, w/o L-Glutamine,

SFM-SUT100

100ml

AK-CHO sus SFM Base w/o Sodium Bicarbonate AK-CHO supp

SFM Supplement 100X Concentrate

SFM-SUT500 SFM-CHS005

SFM-CHS050 SFM-SUT100

500ml 5ml

50ml

100ml

For instructions of Method of Use please refer to Ak-Multicell Method of Use separate page.

05

AUXILIARY SOLUTIONS

AK-Fibronectin - Bovine Fibronectin Solution AK-Fibronectin is an attachment factor that facilitates the attachment and cytoplasmic spreading of anchorage dependent cells. AK-Fibronectin is particularly useful for culturing cells that are incapable of synthesizing their own biomatrix, or when culturing cells in serum-free medium.

PRODUCT

Catalogue #

Size

Fibronectin Solution (Bovine)

SFA-FIB001

1ml

PRODUCT

Catalogue #

Size

Crystalline Trypsin Solution w/o Phenol Red

SFA-TRP100

100ml

SFA-STI020

20ml

AK-Fibronectin

Concentration: 1mg/ml

SFA-FIB005

5ml

Stability

AK-Fibronectin solution is stable for 2 years when stored at 2-8°C. Suggested Coating Procedures

The AK-Fibronectin should be added to the growth medium in the growth vessel, which is then placed in an incubator for 3060 minutes before seeding. The recommended concentration of the AK-Fibronectin is 5 µg/ml of medium.

When the medium is replaced in the days following initial seeding, no further AK-Fibronectin supplementation is required.

AK-Trypsin Solution & AK-SoyTI Solution The use of crystalline trypsin, rather than crude trypsin is often essential for successful, long-term growth of cells in serum-free media. Furthermore, excess Crystalline Trypsin (AK-Trypsin) can be neutralized with Soy Bean Trypsin Inhibitor (AK-SoyTI), thus avoiding the use of serum for this purpose.

AK-Trypsin

AK-SoyTI

Soybean Trypsin Inhibitor

SFA-TRP500

500ml

AK-Freez - Serum-free cell freezing medium Serum-free cell freezing medium is used to cryopreserve mammalian cells, particularly when preserving cells cultured in serum-free media. Akron Biotech’s AK-Freez, serum-free cell freezing medium, contains methylcellulose and DMSO, providing a high percentage of viable cells after freezing and thawing. Cells also show excellent attachment ability and growth performance. Our studies have shown higher viability and adhesion percentages using AK-Freez compared with serum-containing freezing medium. The use of AKFreez is therefore also recommended for cell culture employing serum-supplemented growth media.

Typical Experimental Results % Viability

Stability

Cell line

AK-Freez

3T3 BGM VERO HEp-2 BSC-1

85 91 66 75 82

06

83 83 71 69 77

AK-Freez

100 88 62 100 22

Serum containing AK-Freez

83 88 33 92 10

PRODUCT

Catalogue #

Size

Serum-Free Cell Freezing Medium

SFM-FRZ020

20ml

AK-Freez

AK-Freez is stable for 15 months when stored at 2-8°C.

Serum containing AK-Freez

% Adhesion

INSECT MEDIA

AK-Insect AK-Insect is a serum-free medium optimized for the culture of lepidopteran insect cells. The medium supports both suspension and stationary cultures of Sf-9 cells derived from the pupal ovarian tissue of Spodoptera frugiperda.

Sf-9 cells are suitable hosts for the replication of the baculovirus Autographa colifornica nuclear polyhedrosis virus. This virus, isolated from the Alfalfa looper, is used for the recombinant expression of heterologous proteins in the baculovirus expression vector system (BEVS). Insect cells, infected with this virus, display accumulations of the highly expressed protein polyhedrin, within the nuclea (polyhedral).

The use of AK-Insect for the growth of Sf-9 cells has shown significantly better results than those obtained using TNMFH medium (supplemented Graceâ&#x20AC;&#x2122;s) with 10% fetal bovine serum.

AK-Insect has also shown excellent performance when cultivating high-V cells and in the production of recombinant beta-galactosidase.

Stability

AK-Insect is stable for 12 months when stored at 2-8°C. PRODUCT

Catalogue #

Size

AK-Insect

SFM-INS100

100ml

with L-Glutamine

07

SFM-INS500

500ml

MONONUCLEAR MEDIA

AK-Cell AK-Cell has been developed specifically for use with human mononuclear cells (lymphocytes and monocytes) from peripheral blood. In work with these cells and their sub-populations, it is critical to optimize and define the media formulation as well as pH and temperature. In most cases, these cells are grown in conventional media, supplemented with human serum (A, AB) or fetal bovine serum. However, the use of serum suffers from the following disadvantages: • The presence of non-specific growth factors that interfere with complete activation in the desired direction.

• Inhibitors that limit activation of lymphocytes.

• Lot to lot variation.

• Pathogens introduced via the serum.

The evaluation of antigenic reactions, such as the quantity of the lymphokines generated, and the reaction of the lymphokines to hormones and the growth factors, are all more accurate in the absence of serum.

PRODUCT

Catalogue #

w/o L-Glutamine

SFM-CEL100

AK-Cell

08

SFM-CEL500

Size 100ml

500ml

Applications

Protocols

The applications for the use of AK-Cell are numerous and include:

Following are several examples of the evaluation protocols by which AK-Cell was selected:

• Activation of mononuclear cells with the aid of various mitogens (PHA, CON.A, OKT-3)

• Mitogenic Activation of Mononuclear Cells: Activation was evaluated with different mitogens such as PHA, CON.A and OKT-3. Proliferation was measured by the uptake of radioactive thymidine. The mitogens were added in varying concentrations and thymidine uptake was determined over several days, in order to fully evaluate the specific medium formulation.

• Activation of mononuclear cells with lymphoid cells (RAJI, PEER, BA, MOLT-4, JURKAT) • Production of IL-2 and IL-3 from mononuclear cells

• Activation of Mononuclear Cells with Lymphoid Cells: The activation of the mononuclear cells was carried out using lymphoid cells of various kinds, such as: JURKAT, RAJI, MOLT-4, and BA. Varying ratios between the tumor cells and the mononuclear cells were examined, and the proliferation was measured by uptake of radioactive thymidine.

• Long-term culture of mononuclear cells after activation • Activation of mononuclear cells with interleukin-2 to generate LAK or TIL cells • Activation of mononuclear cells to generate natural killer cells (NK)

• Production of Lymphokines by Activated Mononuclear Cells: The levels of the lymphokines IL-2 and IL-3 were measured in the culture of the mononuclear cells after activation with various mitogens. IL-2 production was measured with the help of the CTLL-2 cell line. These are cytotoxic T-cells from mice, which grow only in the presence of IL-2 in the culture medium.

• Activation of mononuclear cells to generate cytotoxic T cells • Activation of macrophages

• Cytotoxicity: Mononuclear cells were seeded at a concentration of 10(6) cells per ml together with RAJI cells which had been treated with mitomycin C.

• Research on the influence of various cytokines on the production of sub-populations of mononuclear cells • Proliferation of the HIV virus

Varying ratios of the two cell types were examined. At the conclusion of the activation (5-7 days), the lymphocytes were collected, centrifuged, suspended in medium and seeded in microwells to measure proliferation and cytotoxicity. RAJI cells were labeled with radioactive chromium (10 ci in a volume of 0.2 ml), washed three times, suspended at a concentration of 10(5) cells per ml, and divided into microwells containing the above activated lymphocytes.

After 18 hours incubation, the cytolytic activity was evaluated by measuring the radioactive chromium released from the target (RAJI) cells.

• Proliferation of retroviruses in T cells for the purposes of vaccine development Stability

AK-Cell is stable for 12 months when stored at 2-8°C.

09

CULTURED DISHES

Coated with Extracellular Matrix & Serum-Free Medium

AK-ECM The AK-ECM is similar in its chemical composition and organization to naturally occurring basement membranes upon which cells migrate, proliferate, and differentiate in vivo.

Thus, disposable plastic ware coated with ECM mimics the in vivo environment under in vitro experimental conditions.

• Increasing yields: In many instances the minute quantities of materials secreted by cultured cells is a major drawback in the production of cellular materials. Use of serum-free medium presents exciting possibilities for increasing the yield of various cellular products through batch processing methods.

Cells placed in contact with ECM act in the following manner: • Attach rapidly.

• Exhibit high plating and cloning efficiencies.

• Proliferate rapidly.

• Reach a high saturation density.

• Exhibit lower requirements for serum and added growth factors.

• Respond better to physiologically occurring hormones.

• Express differentiated functions.

• Have longer life span.

• Undergo flattening and morphological changes.

• Have better plating consistency.

The Production Line

Akron Biotech offers a complete line of AK-ECM-coated disposable plastic ware. The product line is divided into several groups to assure coverage of the types of disposable plastic ware most commonly used in research laboratories studying in vitro characteristics of mammalian cells.

ECM-coated dishes with serum-free media support the maintenance and normal function of hormone secreting cells such as pancreatic islet cells, hepatocytes pituitary cells, granulosa cells, etc. The ability to culture cells from endocrine organs makes it possible to study control mechanisms of hormone production.

AK-ECM Extracellular MatrixCoated Plastic Ware

The AK-ECM/serum-free medium combination promotes research possibilities on various cellular products due to the following multiple effects:

Tissue Culture Dishes 35mm

ECM-TCD035

Tissue Culture Dishes 90mm

ECM-TCD090

Tissue Culture Dishes 60mm

• Inducing differentiation: The development of appropriate conditions for the maintenance of differentiated cells in culture is a key to studying specific functions of hormone secreting cells and for increasing the yield of secreted products.

Tissue Culture Flasks 25cm Tissue Culture Flasks 80cm

Microtiter 96-Well Plate (flat) 4-Well Plate

• Suppressing fibroblasts: Serum-free media have been shown to suppress the growth of fibroblasts and hence allow the maintenance of almost pure epithelial cell cultures.

6-Well Plate

12-Well Plate 24-Well Plate

Coverslips (round) 22mm

• Purification: Since AK-ECM allows the growth of cells in serum-free media, it facilitates the purification of various cellular products from medium lacking most macromolecular contaminants.

Eight 12mm Filters in 24-Well Plate Four 12mm Filters in 6-Well Plate Four 13mm Filters in 4-Well Plate

10

Catalogue #

ECM-TCD060 ECM-TCF025 ECM-TCF080

ECM-MIT096

ECM-WLP004 ECM-WLP006 ECM-WLP012 ECM-WLP024

ECM-COS022 ECM-FIL024

ECM-FIL006 ECM-FIL004

#units/ package 5 5 5 5 5 1 1 1 1 1 5 1 1 1

INDEX page

3

SFM for Mammalian Cell Culture in Suspension AK-Media hi, AK-Media mid, AK-Media lo AK-Grow hi, AK-Grow lo

4-5

SFM for Adherent and Suspension Cultures AK-CHO adh, AK-CHO sus & AK-CHO supp AK-MultiCell

6

AUXILIARY SOLUTIONS

AK-Fibronectin - Bovine Fibronectin Solution AK-Trypsin Solution & AK-SoyTI Solution

AK-Freez - Serum-Free Cell Freezing Medium

7 8-9 10

INSECT MEDIA AK-Insect

MONONUCLEAR MEDIA AK-Cell

CULTURED DISHES Coated with Extracellular Matrix & Serum-Free Medium AK-ECM