CUSTOM ANTIBODY SERVICES & PRODUCTS

Transcription

1 CUSTOM ANTIBODY SERVICES & PRODUCTS POLYCLONAL SERVICES MONOCLONAL SERVICES ANTIBODY SUPPORT SERVICES ANTIGEN SUBMISSION ANTIBODY PROJECT INITIATION CUSTOM ANTIBODY SERVICES & PRODUCTS

2 CUSTOM ANTIBODY SERVICES & PRODUCTS Section Contents POLYCLONAL SERVICES ANTIGEN SUBMISSION MONOCLONAL SERVICES ANTIBODY PROJECT INITIATION THE CELL CULTURE BAG ZEBRAFISH TERATOLOGY ANTIBODY SUPPORT SERVICES PROJECT INITIATION FORMS End of Section POLYCLONAL ANTIBODY SERVICES Biological Laboratories has been producing high-quality antibodies since 1977, offering a variety of Polyclonal Antibody Services to fit your research and production needs. Each project begins with a free antigen design analysis with 's scientific staff. We offer cost-effective, standard programs, as well as custom protocols designed by our team of professionals to meet your specific needs. Our standard protocols are described in the following pages and have been used with great success for a wide variety of antigen types. If a custom protocol is more appropriate for your project, please contact our Customer Support Staff at or lampire@lampire.com for details on how to get started. EXPRESS-LINE Services With complete packages beginning at just $ per animal, 's EXPRESS-LINE Services are designed for rapid antibody production at a low cost. The 50-day immunization protocol has proven successful for many antigen types and may be extended by selecting the Plus or Extended Services. CLASSIC-LINE Services 's CLASSIC-LINE Services are designed to be flexible to meet the needs of academic and industry scientists alike. The 98-day immunization protocol is ideal for hapten and bacterial antigens, as well as for long-term projects requiring large volumes of antibody. For manufacturing-scale quantities of antisera, we offer the CLASSIC-LINE Extended Service, which moves your animals to a monthly rotation at Day 98 for on-going production. HARMONY-LINE Services HARMONY-LINE Services couple peptide synthesis with antibody development. We offer free consultation on antigen design and will custom synthesize a peptide to your specifications. Peptides are purified to >75% or >85% purity and undergo Quality Control by HPLC and Mass Spec. We offer several pre-packaged programs for two animals using a 57-day protocol, and offer an Extended protocol for larger volumes of antisera. Antibody Support Services A variety of support services are available such as peptide synthesis, antigen design consultation, antigen conjugation, ELISA testing, antibody purification, antibody labeling, and more. Descriptions of these services begin on page 58. Animal Selection Table Use this table to help select the appropriate species for your polyclonal project. Species: Comments: Antigen Required: Test Bleed Volume: Production Volume: Exsanguination Bleed Volume: Rabbit Goat/Sheep Chicken Llama Recommended for research Recommended for production Recommended for highly Recommended for research & volumes of antibody. volumes of antibody. conserved mammalian antigens production volumes of antibody mg/injection mg/injection mg/injection mg/injection 2 ml (approx.) 3-5 ml 2 ml (approx.) 3-5 ml ml/bleed ml/bleed 0-7 eggs/week ml/bleed ml ml N/A ml POLYCLONAL ANTIBODY SERVICES 2009 Biological Labs, Inc. 49

3 CLIENT REPORTS & SAMPLE DATA Monthly Client Boarding Report Each month, clients receive reports similar to those depicted below. These reports detail items such as past and future project milestones and antigen inventories. In addition to understanding the progression of your projects, these reports help ensure the status of individual animals on each project is being tracked. We have found this helps minimize the number of project animals that are pending or awaiting instructions. In addition to monthly client reports, up-to-date immunization protocols can be made available, allowing you to track the status of a project and verify any desired protocol modifications. ELISA Testing As an optional service, will perform ELISA testing of your antisera or purified antibody. Assays can be run against the host antigen or any additional target antigens provided. Upon completion of the assay, 's technical staff will contact you and discuss the results and available options. These may range from protocol and/or antigen modification to the termination of non-responding animals. Biological Laboratories Monthly Client ELISA Data: Example Report Day 30 TB Day 50 PB Feb Prod. March Prod. April Prod. Animal Titer Titer Titer Titer Titer Species ID 1:X 1:X 1:X 1:X 1:X Biological Laboratories Monthly Client ELISA Titer Graph: Example Report Rabbit 687 < Rabbit 688 < Rabbit 689 < Rabbit 690 < Rabbit 691 <20 180** < POLYCLONAL ANTIBODY SERVICES Absorbance (nm) Rabbit Titers: Production Bleed 4/3/09 Client: Contact: Company Scientist CONSOLIDATED CLIENT BOARDING REPORT FOR THE MONTH ENDING JANUARY 31, 2009 P.O. Invest. Project Start Animal Number Term 06 JAN Boost Bleed Dr. Smith 143-AC 01/02/09 Rabbit Feb 8 Feb Dr. Smith 143-AC 01/02/09 Rabbit Feb 8 Feb Dr. Smith 143-AC 01/02/09 Rabbit Feb 8 Feb 22 P.O. Subtotal Dr. Smith 558-NT 10/19/08 Rabbit Awaiting Instructions Dr. Smith 558-NT 10/19/08 Rabbit Awaiting Instructions Dr. Smith 558-NT 10/19/08 Rabbit Awaiting Instructions 3 P.O. Subtotal CX Dr. Smith 08/15/08 Rabbit /26/09 Terminated Dilution Dr. Smith 981-CX 08/15/08 Rabbit /26/09 Terminated Dr. Smith 981-CX 08/15/08 Rabbit /26/09 Terminated A C D y=(a-d)/(1+(x/c)^b)+d: B R^2 P.O. Subtotal 0 Rabbit 687 (R687 april: Concentration vs Meanvalue) Total Rabbits 6 Rabbit 688 (R688 april: Concentration vs Meanvalue) Rabbit 689 (R689 april: Concentration vs Meanvalue) Rabbit 690 (R690 april: Concentration vs Meanvalue) Rabbit 691 (R691 april: Concentration vs Meanvalue)

4 EXPRESS-LINE SERVICES The EXPRESS-LINE Services are 's most cost-effective programs and are recommended for most antigen types. The Basic Service is quickest, yielding antibody in just 50 days. The Plus Service extends the program for one week and yields an additional amount of antibody. The Extended Service is your option to indefinitely continue your program, delivering antibody to you on a long-term basis. Basic Service Day Procedure: volumes in ( ) are for rats 1 Pre-bleed. (200ul) 2 ml serum per other animals. Inject (150ul) 0.5 ml antigen mixed with (150ul) 0.5 ml Complete Freund's adjuvant. 21 Inject (150ul) 0.5 ml antigen mixed with (150ul) 0.5 ml Incomplete Freund's adjuvant. 30 Test bleed. Sample shipped for titer testing/screening. 42 Inject (150ul) 0.5 ml antigen mixed with (150ul) 0.5 ml Incomplete Freund's adjuvant. 50 Rabbits, Goats, Sheep, Donkeys, Llamas, Rats: Production bleed. Chickens: Egg Collection. Project terminated. Pricing Schedule Rabbit Goat/Sheep Chicken Donkey/Llama Rat Project Initiation Fee: $ $ $ $ $ Board/Maintenance (50 days): $ $ $ $ $ Production Bleed: No Charge $ * No Charge $ * $ Project Termination Fee: $ $ $ $ $ Total Price per Animal: $ $ $ $ $ Plus Service - The above 50-day Basic protocol, plus the services below. Day Procedure Chickens: Eggs collected daily. Project terminated. 57 Rabbits, Goats, Sheep, Donkeys, Llamas, Rats: Exsanguination bleed. Project terminated. Pricing Schedule Rabbit Goat/Sheep Chicken Donkey/Llama Rat Project Initiation Fee: $ $ $ $ $ Board/Maintenance (57 days): $ $ $ $ $ Production Bleed: No Charge $ * No Charge $ * $ Egg Collection: N/A N/A No Charge N/A N/A Exsanguination Bleed: $ 55.00** $ ** $ $ ** $ Project Termination Fee: $ $ $ $ $ Total Price per Animal: $ $ $ $ $ Extended Service - The above 50-day Basic protocol, plus the services below. (Additional antigen may be required.) Week A B C D Procedure: volumes in ( ) are for rats Rabbits, Goats, Sheep, Donkeys, Llamas, Rats: Production bleed. Chickens: Test bleed. Daily egg collection continues. Rest. Injection. Boost (150ul) 0.5 ml antigen mixed with (150ul) 0.5 ml Incomplete Freund's adjuvant. Rest. Thereafter, repeat monthly rotation as above. Pricing Schedule Rabbit Goat/Sheep Chicken Donkey/Llama Rat First 50 Days: $ $ $ $ $ Board/Maintenance (per day): $ 2.50 $ 3.25 $ 2.50 $ 7.75 $ 2.25 Injection (1 per month): No Charge No Charge No Charge No Charge No Charge Production Bleed (1 per month): No Charge $ 0.50/ml N/A $ 0.50/ml No Charge Egg Collection: N/A N/A No Charge N/A N/A Production Bleed (2nd bleed in month): $ $ 0.50/ml $ $ 0.50/ml $ Project Termination Fee: $ $ $ $ $ Total Price per Animal: All totals will vary, based on project length. POLYCLONAL ANTIBODY SERVICES *Price is an estimate based on a Production Bleed volume of 300 ml for goats/sheep and 1 L for donkeys/llamas at $0.50/ml. **Price is an estimate based on Exsanguination Bleed volumes of 70 ml for rabbits, 700 ml for goats/sheep and 2.5 L for donkeys/llamas at $0.50/ml. Note: Llama volume may have a larger variation depending on total weight of the animal. 51

5 CLASSIC-LINE SERVICES The CLASSIC-LINE Services are designed to optimize antibody production against small, non-immunogenic molecules such as haptens. The Basic Service yields antibody in 98 days and the Extended Service allows you to continue your project as long as needed to obtain the amount of antibody desired. Basic Service Day Procedure 1 Pre-bleed. 2 ml serum per animal. Pre-immune serum shipped with first test bleed. Inject 0.5 ml antigen mixed with 0.5 ml Complete Freund's adjuvant. 7 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant. 14 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant Rest. 28 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant Rest. 42 Test bleed. Chickens: Daily egg collection begins Rest. 56 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant Rest. 70 Test bleed. [Note: A production bleed may be scheduled here if the animal has achieved sufficient titer.] Rest. 84 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant Rabbits, Goats, Sheep, Donkeys, Llamas: Rest. 98 Rabbits, Goats, Sheep, Donkeys, Llamas: Production bleed. Chickens: Egg collection. Project terminated. Pricing Schedule Rabbit Goat/Sheep Chicken Donkey/Llama Project Initiation Fee: $ $ $ $ Board/Maintenance (98 days): $ $ $ $ Production Bleed: No Charge $ * No Charge $ Project Termination Fee: $ $ $ $ Total Price per Animal: $ $ $ $ Note: If an exsanguination bleed is desired, it must be requested separately in writing and is invoiced at: For rabbits & chickens $55. For goats, sheep, donkeys and llamas $0.50/mL of serum. POLYCLONAL ANTIBODY SERVICES *Price is an estimate based on a Production Bleed volume of 300 ml for goats/sheep and 1 L for donkeys/llamas at $0.50/ml. Note: Llama volume may have a larger variation depending on total weight of the animal. Extended Service - The 98-day Basic protocol, then (Note: Requires additional antigen) Week A B C D Procedure Rabbits, Goats, Sheep, Donkeys, Llamas: Production bleed. Chickens: Test bleed. Daily egg collection continues. Rest. Injection. Boost 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant. Rest. Thereafter, repeat monthly rotation as above. Pricing Schedule Rabbit Goat/Sheep Chicken Donkey/Llama First 98 Days: $ $ $ $ Board/Maintenance (per day): $ 2.50 $ 3.25 $ 2.50 $ 7.75 Injection (1 per month): No Charge No Charge No Charge No Charge Production Bleed (1 per month): No Charge $ 0.50/ml N/A $ 0.50/ml Egg Collection: N/A N/A No Charge N/A Production Bleed (2nd bleed in month): $ $ 0.50/ml $ $ 0.50/ml Project Termination Fee: $ $ $ $ Total Price per Animal: All totals will vary based on project length 52

6 HARMONY-LINE SERVICES The HARMONY-LINE Services combine peptide synthesis and polyclonal antibody development into a low package price. The ECONOMY Service is the most cost-effective program, using a >75% pure peptide and yields antibody in just 50 days. The prices listed for the Basic and Extended Services described below involve a >85% pure peptide, but each of these services may substitute a >75% pure peptide upon your request. Add ELISA, Western Blot, purification, or conjugation to your project. Check out our Antibody Support Services. Economy Service: >75% Pure Peptide (up to 20 a.a.) Rapid, 50-Day Protocol Conjugation to KLH One Production Bleed per Animal (Egg Collection for Chickens) Day Procedure 1 Pre-bleed. 2 ml serum per animal. Pre-immune serum shipped with first test bleed. Inject 0.5 ml antigen mixed with 0.5 ml Complete Freund's adjuvant. 21 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant. 30 Test bleed. Sample shipped for titer testing/screening. 42 Inject 0.5 ml antigen mixed with 0.5 ml Incomplete Freund's adjuvant Chickens: Eggs collected. 50 Rabbits, Goats, Sheep, Donkeys, Llamas: Production bleed. Pricing Schedule 2 Rabbits 2 Goats/Sheep 2 Chickens Package Price $ $ $ Includes: KLH-Conjugated Peptide (75% pure) 5-10 mg 5-10 mg 5-10 mg Unconjugated Peptide mg mg mg Test Bleed 2 ml 5 ml 2 ml Production Bleed/Egg Collection ml ml 5-7 eggs Basic Service: The above 50-day Basic protocol with a >85% Pure Peptide (up to 20 a.a.) Pricing Schedule 2 Rabbits 2 Goats/Sheep 2 Chickens Package Price $ $ $ Includes: KLH-Conjugated Peptide (85% pure) 5-10 mg 5-10 mg 5-10 mg Unconjugated Peptide mg mg mg Test Bleed 2 ml 5 ml 2 ml Production Bleed/Egg Collection ml ml 5-7 eggs Plus Service: Add an Exsanguination Bleed to either the Economy or Basic Services Day Procedure Chickens: Daily egg collection. 57 Rabbits, Goats, Sheep, Donkeys, Llamas: Exsanguination bleed, project terminated. Chickens: Project terminated. Pricing Schedule 2 Rabbits 2 Goats/Sheep 2 Chickens Additional Price $ $ $34.75 (add to Economy and Basic prices) Includes: Exsanguination Bleed/Egg Collection ml ml eggs Extended Service: The above 50-day Basic protocol, plus the services below. (Additional antigen may be required.) Week A B C D Procedure Rest. Injection. Boost 0.5 ml antigen mixed with 0.5ml Incomplete Freund s Adjuvant. Rest. Rabbits, Goats, Sheep, Donkeys, Llamas: Production bleed. Chickens: Daily egg collection. POLYCLONAL ANTIBODY SERVICES Pricing Schedule 2 Rabbits 2 Goats/Sheep 2 Chickens 30-Day Period $ $ $ Includes: Production Bleed/Egg Collection ml ml eggs 53

7 MONOCLONAL SERVICES OVERVIEW 's Biotechnology Research and Resource Center in Pipersville, PA, offers an array of services for the research, development and production of monoclonal antibodies, using both in vivo and in vitro culture methods. Antibody production services are conducted under cgmp guidelines. is FDA Registered, USDA Licensed, AAALAC accredited, and holds an NIH Assurance. All services are kept completely confidential. Our trained staff are a valuable resource available to you to help refine the details of the scope and design of your project. MONOCLONAL ANTIBODY SERVICES DISCOVERY-LINE Services Designed to meet the needs of academic and industry scientists alike, 's Discovery-Line Services provide hybridoma development at an affordable price. Each project begins with a thorough biochemical and biological analysis of your antigen, then a protocol is developed for your review by means of a Project Scope & Definition Form. After protocol approval immunizations begin, followed by titer screening and collaborative data review. Fusions and cloning then ensue as appropriate. At the end of the project the resultant cell line is delivered to, and is the property of, the client. As a safety measure can hold, store, and maintain daughter cell lines, which are available to the client at any time. (See page 60, Cell Bank Storage, for a description of services and fees.) SELECT-LINE Services 's Select-Line Services are cost-effective solutions for small- or large-scale antibody production of established cell lines. Performed at the Biotechnology Research and Resource Center in Pipersville, PA, a variety of cell culture techniques are available, featuring the Cell Culture Bag (page 57). Additional production services include roller bottles, bioreactors, and murine ascites. 's standard medium is a DMEM/F12 derivative; use of other media may incur an additional charge. Note: Both the Discovery-Line and Select-Line Services use Balb/C mice; other strains may incur an additional charge. 54

8 DISCOVERY-LINE SERVICES High Quality Hybridoma Development and Characterization 's Discovery-Line Services offer high-quality hybridoma development at a very affordable price. Designed to meet the needs of both industry and academic scientists, each project begins with free Antigen Design and Project Scope & Definition meetings. Projects are divided into three phases, each with its own billing cycle to spread your costs out over the course of the project. A description of each phase is presented below. Phase I: Project Design and Immunization Each project begins with a Scope of Project meeting between you and our technical staff in an effort to assure the success of your project. Mice or rats are then immunized according to the schedule below and titer is monitored by ELISA until sufficient titer is reached to warrant fusion between immunized animal spleen cells and an immortal, non-secreting myeloma cell line. Day Procedure 1 Pre-bleed and first injection. 28 Second injection. 38 First test bleed. Note: Subsequent injections and test bleeds, if required, will be performed only after consultation with our technical staff. Pricing Schedule No. of Animals Cat. No. Price Project Design and Immunization 5-Mice $1, Mice $1,650 3-Rats $2,200 Phase II: Cell Fusion After candidate animals are selected, the number of required fusion plates is determined, and the fusions are performed. Fusion plates are screened by ELISA to identify hybrid cell lines that produce antigen specific antibody. Several fusions may be required to obtain a hybrid cell line that produces the desired antibody. Fusion candidates will be approved by the client before the fusions are performed and charges are incurred. Pricing Schedule Desc Cat. No. Price Cell Fusion Mouse - Single Antigen $4,125 Mouse - Dual Antigen $4,675 Rat - Single Antigen $4,950 Rat - DualAntigen $5,500 Phase III: Cloning Positive cell lines from Phase II will be expanded to produce sufficient culture to preserve the parent cell line and to subclone the identified positives by limiting dilution. Typically 3 cell lines are chosen for Phase III, but this number can be increased based on client requirements. The expansion culture of the selected positive cell lines will be preserved in liquid nitrogen. Up to fifteen (15) additional positive cell lines may be cryopreserved individually at or up to 100 may be cryopreserved as pools. These cell lines may also be shipped to you if requested. (See page 60, Cell Bank Storage, for a description of services and fees.) Recloning of selected positives will be performed as needed to achieve antibody production stability. Clones must exhibit >95% antigen-specific antibody production to be considered stable. Pricing Schedule Desc Cat. No. Price Cloning Mouse - Per Clone Per round of Subclone - 1 Antigen $330 Mouse - Per Clone Per round of Subclone - 2 Antigen $495 Rat - Per Clone Per round of Subclone - 1 Antigen $440 Rat - Per Clone Per round of Subclone - 2 Antigen $605 MONOCLONAL ANTIBODY SERVICES 55

9 Cost-Effective, Customizable, Cell Culture Production for Established Hybridoma Cell Lines 's Select-Line Services enable small- and large-scale antibody production for established hybridoma cell lines. A variety of cell culture techniques are available, featuring the new Cell Culture Bag. Additional services include ascites, roller bottle, and bioreactor production. Prices include the use of 's standard medium. A medium of your choice may be substituted, but may incur an additional charge. Pricing Schedule Culture Bag Production Please Inquire Ascites production employs Balb/C mice, with ascites fluid collected and pooled at the end of the project. Each mouse yields approximately 2-5 ml ascites. Alternate strains of mouse may be used, but may incur an additional charge. Production volume is approximately 2-5 ml ascites per mouse. Prices include husbandry of mice for up to 35 days, with 1 injection of cells and up to 3 taps. Pricing Schedule Cell Line Expansion Please Inquire Note: AAALAC regulatory guidelines require written assurance that the proposed laboratory animal use is scientifically justified and that in-vitro or other methods have been evaluated and found unsuitable. MONOCLONAL ANTIBODY SERVICES Roller bottles are a cost-effective alternative to large-scale antibody production using murine ascites. Typical lot sizes are 1-10 g of antibody. Production volume is approximately mg/liter of roller bottle culture for an average producing cell line. Prices include the use of 's standard media; media of your choice may be substituted, but may incur an additional charge. Pricing Schedule Roller Bottle Production Note: Bulk discounts are available. Please inquire. Please Inquire Bioreactor formats include hollow fiber, stirred tank and microcarrier systems. Single-use bioreactor technologies are offered for GMP-compliant procedures. Hollow fiber systems are ideal for non-adherent cell lines and work well with most cell lines tested. Production volumes of up to 300 mg per day are possible and the system can be used to grow and harvest cells and cell products other than antibody. Stirred tank systems are useful for both adherent and non-adherent cell lines and are capable of producing 1-3 g antibody per week. Microcarrier systems employ packed and fluidized bed techniques to increase the surface area for cell attachment in stirred tank bioreactors. Production volumes of mg per day are possible. Prices include the use of s standard medium; both the substitution with a medium of your choice or the inclusion of a medium optimization strategy are available, but may incur an additional charge. Pricing Schedule Bioreactor Production Please Inquire 56

10 The OMNI C3 CELL CULTURE BAG AN ELEGANTLY SIMPLE APPROACH TO GROWING CELLS EFFICIENT, ECONOMICAL AND ENVIRONMENTALLY RESPONSIBLE... The Cell Culture Bags are an efficient, economical and environmentally responsible cell culture alternative to many other traditional systems such as flasks and roller bottles. These microscopicallytransparent, single-use bags facilitate the handling and expansion of many different cell culture types from standard hybridoma cells/antibody production to customized cell propagation and recombinant protein production. Ranging in size from 50ml up to 2 liter, the presterilized bags can be filled to the full capacity for higher productivity per incubator space. The Cell Culture Bags are equipped with a wide access port for easy media delivery, sampling and harvesting. The high gas permeability of the bag material maximizes cell growth and yields over extended growth periods. Cell Culture Bags are easy to use from start to finish-- easy to fill, easy to observe cell growth, easy to maintain, and easy disposal. HIGH PRODUCTIVITY HIGH GAS PERMEABILITY Free exchange of O 2 and CO 2 allow enhanced cell respiration and elimination. INCREASED CELL COUNTS Growing more cells in fewer containers reduces cost and labor. INCREASED ANTIBODY YIELDS You ll experience fast growth rate and extended cell viability. FULL CELL CULTURE BAG VOLUME UTILIZED More cells per volume and less incubator space required compared to roller bottles or T flasks. SIMPLIFIED PROTOCOLS MINIMAL HANDLING Minimizes chances for risk of contamination. NO EXPENSIVE EQUIPMENT NEEDED Only a basic CO 2 incubator is required. NO ADDITIVES REQUIRED No cell feeding or trypsin required for cell extraction. A simpler process reduces the margin of error and the chance of contamination in the lab. EASY TO USE PRESTERILIZED Individually packaged bags are ready to use. WIDE ACCESSIBLE PORT Convenient for media delivery and cell retrieval. CLEAR MICROSCOPIC VIEWING Effective cell observation without the need to open the bag to obtain a sample. MINIMAL STORAGE & DISPOSAL SPACE REQUIRED The Cell Culture Bag collapses to a flat profile, taking up far less storage and disposal space than alternative cell culture vessels. EASY ON THE ENVIRONMENT ENVIRONMENTALLY FRIENDLY More cells in fewer, and more compact containers, plus condensed waste removal make the Cell Culture Bag more environmentally efficient. Ordering Information Description Cat. No. Price 50-ml Fill Volume (15 per box) $ ml Fill Volume (10 per box) $ Liter Fill Volume (5 per box) $ Liter Fill Volume (5 per box) $ 215 The CELL CULTURE BAG 57

11 ANTIBODY SUPPORT SERVICES Support Services for Polyclonal and Monoclonal Antibodies has provided quality antibody support services since From antigen design and peptide synthesis to antibody purification and labeling, our expert staff is available to assist you with every phase of antibody development. The success of your antibody project is very important to us. As such, we encourage you to take advantage of our free Antigen Design and Protocol Review described below. Peptide Synthesis Enhance your monoclonal and polyclonal antibody development with peptide synthesis. Included as part of our HARMONY-LINE Services (page 53), peptides are carefully designed and synthesized under the highest quality standards. Quality Control measures include Mass Spec and HPLC. Peptide modifications are also available. Purity Levels: >75% >85% >90% >95% Modifications: Acetylation Nitrosylation Phosphorylation Peptides are synthesized on a custom basis; please contact our Customer Support Staff for a price quote. Antigen Design and Protocol Review Prior to beginning any monoclonal or polyclonal development project, you have the option of meeting with 's scientific staff for consultation on antigen design and immunization protocol. Using computer-aided technology, scientists will analyze peptide sequences relevant to your target antigen and review with you the predicted immunogenicity. This service is provided free of charge and aids us in selecting the appropriate immunization protocol for your antigen. While we cannot guarantee every peptide sequence will generate an immune response in the host animal, this service has proven very successful in maximizing the success of each antibody development project. Pricing Schedule Antigen Design Analysis No Charge ANTIBODY SUPPORT SERVICES Antigen Conjugation To improve immunogenicity, certain antigens may require coupling to a larger molecule. These antigens may be coupled to KLH, BSA, or other requested proteins. Coupling methodologies include EDC and Glutaraldehyde. Pricing Schedule KLH Conjugation $275 BSA Conjugation $275 Alkaline Phosphatase Conjugation Inquire Horseradish Peroxidase Conjugation Inquire FITC Conjugation Inquire Biotin Conjugation Inquire Colloidal Gold Conjugation Inquire ELISA Testing ELISA testing can be useful in measuring and tracking the concentration of antibody in culture supernatants, bleeds, or purified materials. Pricing Schedule Plate Charge (up to 4 samples) $99 Note: Cost per project will vary based on the frequency of testing and type of ELISA. Also, additional antigen may be required. 58

12 offers several services to help evaluate the purity, concentration, and specificity of your antibody. Please contact our Customer Support Staff for details of these services. Pricing Schedule BCA Total Protein Assay Bradford Total Protein Assay ELISA Titer Screen HPLC Immunoelectrophoresis Radial Immunodiffusion SDS-PAGE Western Blot $ per sample $ per sample $27.50 per sample ($99 minimum plate charge) $110 per sample $330 per plate $165 per plate (5 samples) $ per gel $385 up to 2 samples offers a variety of purification services to complement your antibody project. From salt fractionation to antigen-specific affinity chromatography, our expert staff will purify your antibody by any of the methods below. Purification Services Delipidation 1 liter: $ > 1 liter: Inquire Ammonium Sulfate Fractionation 1 liter: $275 > 1 liter: Inquire DEAE Chromatography 25 ml: $275 > 25 ml: Inquire IgY Purification Purification of first dozen eggs $330 Each additional dozen eggs $165 Immunoaffinity Chromatography 50 ml $715 >50 ml to 100 ml $1,187 >100 ml to 1000 ml Inquire Note: Pricing is for a single pass, additional passes $265 each >25mL >50mL >100mL >300 >500 25mL to to to to to Protein A Chromatography: 50mL 100mL 300mL 500mL 1000mL Ascites, PBS, w/ or w/o azide Inquire Inquire Inquire Inquire Inquire Inquire Monoclonal Supernatant, PBS w/ or w/o azide $ $ $ $ $ $ Rabbit Serum, PBS w/ or w/o azide $ $ $ $1, $1, $ 2, Protein G Chromatography: Ascites, PBS, w/ or w/o azide Inquire Inquire Inquire Inquire Inquire Inquire Monoclonal Supernatant, PBS w/ or w/o azide $ $ $ $ $ $ 1, Rabbit Serum, PBS w/ or w/o azide $ $ $ $ 1, $ 2, $ 3, Goat Serum, PBS w/ or w/o azide $ $ $1, $ 2, $ 3, $ 6, Note: All pricing is listed using a non-dedicated column. If a dedicated column is required, please inquire for pricing. Pricing for other species available upon request. Purified antibodies can be labeled for use in various applications. Consultation with our scientific staff will assist you in selecting the reporter group and coupling chemistry best suited for your applications. Please inquire for pricing. Reporter Group Alkaline Phosphatase Biotin Colloidal Gold Fluorescein Isothiocyanate (FITC) Horseradish Peroxidase (HRP) Chicken immunoglobulin yield quickly declines unless yolks are stored in buffer. offers the additional service of separating the yolks and storing them in buffer. Yolks treated as such can then be shipped to the client for long-term storage at 2-8 C. ANTIBODY SUPPORT SERVICES Pricing Schedule Processing of One Dozen Yolks $121 59

13 ANTIBODY SUPPORT SERVICES - continued Support Services for Monoclonal Antibodies Cell Line Optimization will take an existing cell line that has been provided and optimize the media formulation for growth and production of antibody. Price is dependent upon the number and type of media formulations to be used for the optimization. Please contact our Customer Support Staff for further information. Adaptation to Serum-Free Culture will take an existing cell line and adapt the cell line from a culture media containing serum to a serum-free media culture. Price is dependent upon the type of media to be used for the adaptation and the original serum content of the media. Please contact our Customer Support Staff for further information. Cell Bank Production can make Master and Working Cell Banks of any size to fit your needs. Cell bank reports will be generated and sent for review. Please contact our Customer Support Staff for services such as storage and annual testing for Viability, Doubling-Time, and Antibody Yield. Related fees are outlined below in Cell Bank Storage. Cell Bank Storage has the capabilities of storing cell banks at our facilities. recommends that all cell banks that have been created, whether by or another contractor, be stored at two different locations for safety purposes. can test the cell banks on a yearly basis as well. will expand 1 vial and determine the viability 24 hours, 48 hours, and 96 hours after thaw, the doubling time and antibody yield with results being reported in a one page summary. ANTIBODY SUPPORT SERVICES Pricing Schedule Set-up Fee: $ Storage Fee: $2.25 per vial per year Annual Fee for Viability, Doubling Time, and Antibody Yield Testing: $550 Sterility and Mycoplasma Testing Sterility testing results are based on 21 CFR or USP XXIII. Generally, 14 days at C in fluid thioglycollate and 14 days at C in Soybean Casein Digest medium. Mycoplasma testing reports via indirect DNA Hoechst Staining and Agar and Broth media isolation of culturable strains of mycoplasma. Pricing Schedule Creation of a 25-Vial Cell Bank with Mycoplasma and Sterility Testing: $3,500 60

14 ANTIGEN SUBMISSION Antigen Types Certain types of antigen may require additional processing or conjugation prior to injection. The following is a list of three general antigen types and the possible additional processing or conjugation that may be required. Ready-to-Inject Antigens: Protein extracts, purified proteins, peptide-protein conjugates, and proteins bound to solid supports (e.g., beads). These molecules usually require no further modification or processing. Antigens Requiring Further Processing: Proteins in gel slices or on nitrocellulose membranes. These antigens require homogenization prior to injection into the host animal and incur a $27.50 processing charge. Please contact our Customer Support Staff for details regarding the submission of these antigens. Antigens From Gel Slices: When submitting protein contained in a gel after electrophoretic separation, the following guidelines should be considered: - Nitrocellulose blots are not recommended for immunization; - The gel should be stained with Coomassie Blue and destained with an acetic acid/methanol wash; - Excise the band of interest and estimate the amount of protein; - Run the highest sample concentration possible to limit the amount of acrylamide injected; - Place the gel slice(s) in a conical tube with a damp gauze or paper strip to prevent the gel from drying out; - Do not ship the gel slice with excess water or buffer; - Ship the gel slice on cold packs by overnight carrier. Antigens Requiring Conjugation: Small peptides or haptens. These antigens must be coupled to a carrier protein such as KLH or BSA prior to injection and incur an additional conjugation charge. (See page 58.) Guidelines for Shipping Antigen All antigen should be shipped in the following manner: 1. Concentrate antigen to twice the working concentration. 2. Label tube with antigen name, concentration, and volume. 3. Place in a leak-proof, sealed tube. 4. Antigen should be shipped as it is to be stored. If frozen, ship on dry ice. (See Special Note below.) If refrigerated, ship on ice packs at 2-8 C. Note: Human sourced antigens must be accompanied by a certificate of negativity for infectious diseases. Also, please include an MSDS or safe handling instructions for each antigen. Special Note for Shipments Containing Dry Ice If antigen must be shipped with dry ice, DO NOT place the dry ice in a sealed container of any type. Pressure may buildup in the sealed container, resulting in a very hazardous condition upon package delivery. If dry ice is used, please make sure the container is not air-tight. ANTIGEN SUBMISSION 61

15 ANTIBODY PROJECT INITIATION How to Initiate a Polyclonal Project 1. Please use the forms provided on the following pages or contact the Customer Support Staff to obtain the following: Polyclonal Antibody Project Initiation Form Conjugation and Peptide Questionnaire (if required) IACUC Questionnaire - Polyclonal Projects 2. Fully complete the Polyclonal Antibody Project Initiation Form, by answering all questions, selecting one of the protocols, and then signing. (See pages for details of the pre-defined protocols available.) Note: If you would like a custom protocol, please contact our Customer Support Staff. 3. When completing the antigen section, please refer to Antigen Types (page 61) to determine whether your antigen requires any additional processing or modifications. 4. Refer to 's Antibody Support Services (pages 58-60) for any additional services that may be beneficial to your project. 5. Fax all signed and completed forms to at or to projman@lampire.com. Note: In addition, please fax/ written confirmation of your Purchase Order with the Project Initiation form. All orders will be confirmed by or fax upon technical review and confirmation of price. We will notify you of the shipping address. How to Initiate a Monoclonal Project 1. Contact to obtain the appropriate forms for your project. Our Customer Support Staff will assist you in selecting from the following: Hybridoma Project Initiation Form or SELECT-LINE Project Initiation Form Conjugation and Peptide Questionnaire (if required) IACUC Questionnaire - Monoclonal Projects ANTIBODY PROJECT INITIATION 2. Fully complete the Hybridoma Project Initiation Form or the SELECT-LINE Project Initiation Form, by answering all questions, selecting one of the protocols, and then signing. 3. When completing the antigen section, please refer to Antigen Types (page 61) to determine whether your antigen requires any additional processing or modifications. 4. Refer to 's Antibody Support Services (pages 58-60) for any additional services that may be beneficial to your project. 5. Fax all signed and completed forms to at or to projman@lampire.com. Note: In addition, please fax/ written confirmation of your Purchase Order with the Project Initiation form. All orders will be confirmed by or fax upon technical review and confirmation of price. We will notify you of the shipping address. 62

16 ZEBRAFISH EMBRYO CULTURE TERATOGENICITY SCREEN Normal Valproic Acid Treatment with Jaw Malformation ZEC another initiative to REDUCE, REFINE & REPLACE The zebrafish (Danio rerio) has become an essential tool in genetic studies, particularly for studying spontaneous and induced mutations. The zebrafish has proven to be a useful animal model for studying teratology, toxicology, drug discovery, ecological and environmental sciences, plus many other areas of biomedical research. When compared to the traditional laboratory animal models (e.g., mice, rats) the zebrafish requires less space for housing, is simpler to maintain, and reproduces far more prolifically (2,000 offspring/year vs offspring/year). Rapid, cost-effective teratogenic liability assessment of discovery st age compounds Compounds administered at same developmental stage as in mammalian teratology studies with morphology assessed at fetal-stage equivalent Superior concordance (87%) for classifying in vivo outcome compared to other in vitro developmental tox assays Formal report and complete data analysis package detailing teratogenic potential and dysmorphology associated with compound-exposed embryos Pricing Schedule: Compounds Compounds Compounds Price per Compound $ 2, $ 2, $ 2, This service is available now and custom quotes are generated after free consultation. For more information contact your Lampire representative, call us at , or us at info@lampire.com. ZEBRAFISH TERATOGENICIT Y SCREEN 63

17 RAT WHOLE EMBRYO CULTURE TERATOGENICITY SCREEN Normal Diphenylhydantoin Treatment with First Arch Malformation rwec meeting the animal use initiative to REDUCE, REFINE & REPLACE Fast, cost-effective assay providing unbiased assessment of teratogenic risk Well-established model for assessing teratogenic properties of test agents Compounds administered and cultured during organogenesis, a critical window of sensitivity to most teratogens Quantitative morphological score system used to assess overall growth and grade the severity of abnormalities Effective assay for many water-insoluble compounds ANTIBODY PROJECT INITIATION FORMS Formal report and complete data analysis package detailing teratogenic potential and dysmorphology associated with compound-exposed embryos Pricing Schedule: Standard 1 Dose Study Price $ 9,700/Study* *Price per study includes a single dose per compound (typically 1 µm) along with a solvent, negative, and positive control performed in duplicate assays. Discounted pricing available when requesting multiple studies. Other custom configurations available upon request. Please contact us for a customized quote. This service is available now and custom quotes are generated after free consultation. For more information contact your Lampire representative, call us at , or us at info@lampire.com. 64

18 Sample rwec Report Sample ZEC Report REPORT SAMPLES 65

19 ANTIBODY PROJECT INITIATION FORMS On the following pages you'll find the forms needed to begin either a Polyclonal or Monoclonal Antibody Project. Use the forms provided on the following pages or contact the Customer Support Staff to obtain the following: Polyclonal Antibody Project Initiation Form Hybridoma Project Initiation Form SELECT-LINE Project Initiation Form Conjugation Questionnaire IACUC Questionnaire - Polyclonal Projects IACUC Questionnaire - Monoclonal Projects Please photocopy the forms on the following pages as needed, then complete, sign, and fax to Editable pdf s can also be downloaded at under Literature and Forms, completed and submitted electronically via . ANTIBODY PROJECT INITIATION FORMS 66

20 Polyclonal Antibody Project Initiation Form 1. General Information Company / Institution: Investigator Name: Contact Name: Phone: Fax: 2. Billing / Shipping Information Accounts Payable contact: Company: Dept./Bldg./Room#: Address: Project Name / Number: PO# Credit Card# (Name on Card + Exp ): Ship to contact: Company: Dept./Bldg./Room#: Address: City, State, Zip City, State, Zip Phone: Carrier (FedEx, UPS, etc.) Fax: Your carrier #: 3. Antigen Information Immunogen name: Format (liquid, powder, etc.) Tag or conjugate name: Screening antigen name: Format (liquid, powder, etc.) Tag or conjugate name: Storage conditions: 4 C -20 C -70 C Storage conditions: 4 C -20 C -70 C No. of containers: No. of containers: Volume per container: Volume per container: Concentration / Dosage Conc: Dose: Concentration: Potentially Biohazardous?* Potentially Biohazardous?* Handling instructions (required if biohazardous): * If attenuated, please provide documentation * 4. Protocol Selection (please check only one) Yes No * Unknown Yes No * Unknown Handling instructions (required if biohazardous): If attenuated, please provide documentation CLASSIC-LINE Basic (98-day protocol) EXPRESS-LINE Basic (50-day protocol) HARMONY-LINE Basic (50-day protocol with optional exsanguination bleed at day 57) OTHER (Please attach protocol) CLASSIC-LINE Extended (98-day protocol with monthly option to extend project) EXPRESS-LINE Plus (50-day protocol with exsanguination bleed at day 57) HARMONY-LINE ELISA (50-day protocol with optional exsanguination bleed at day 57 and one ELISA Screen) EXPRESS-LINE Extended (50-day protocol with monthly option to extend project) HARMONY-LINE Extended (50-day protocol with monthly option to extend project) 5. Species and Number of Animals Rabbit(s) Goat(s) Sheep Chicken(s) Mice Rat(s) Other: Qty: Qty: Qty: Qty: Qty: Qty: Qty: 6. Immunochemistry Services ELISA: Yes No Day 0 and test bleed only All bleeds Other: Purification: Yes No Protein A Protein G Affinity Other: Conjugation: Yes No KLH BSA Other: Peptide Synthesis: Yes No Sequence: SP200 38F1 Rev 2 Page 1 of 2 BIOLOGICAL LABORATORIES Tel: Fax: lampire@lampire.com

21 Polyclonal Antibody Project Initiation Form 7. IACUC (Institutional Animal Care and Use Committee) Requirements Lampire Biological Laboratories, Inc., in compliance with federal regulations, presents each animal-use project to IACUC (Institutional Animal Care and Use Committee) for approval prior to project initiation. Criteria for acceptance of a project are established in 9CFR, Chapter 1: Animal and Plant Health Inspection Service, USDA, Subchapter A Animal Welfare. To expedite initiation of your project, please complete the following questions and assurances as thoroughly as possible. If your company or institution is a USDA-registered facility with an IACUC of its own, Lampire may accept an approved Animal-Use Proposal from that committee. If any questions arise during our IACUC review of your project, a committee member will contact you for clarification. Please briefly state your rationale for the appropriateness of the species of animals used for this activity: (i.e., "Goats are appropriate host animals due to the volume of serum required in our project." or "Sheep are appropriate host animals due to their immunologic response to the antigen refinements. " or "Donkeys are appropriate host animals for this project as the characteristics of their antisera are consistent with established requirements for further manufacturing." Please briefly state your rationale for the number of animals used for this activity: (i.e., "One animal will yield an adequate volume of antiserum for determining an antibody response. or Three animals will yield an adequate volume of antiserum for testing purposes. or Ten animals will yield the necessary volume of antiserum required for manufacturing purposes. Your signature below indicates that this project does not unnecessarily duplicate previous experiments or projects, the number of animals has been minimized and alternatives to animal use have been considered. Client Signature Print Name 8. Labeling and Shipping Instructions Please label your antigen(s) accordingly as depicted below. Improperly labeled antigen(s) may delay the start of your project. IMMUNOGEN Project: Client Antigen: Conjugate: Lot #: Conc: mg/ml Vol: ml Storage: C SCREENING ANTIGEN Project: Client Antigen: Conjugate: Lot #: Conc: mg/ml Vol: ml Storage: C Unless otherwise instructed by project management, ship antigen and a copy of this completed form to: 9. Project Approval ( use only) Print a copy of the completed form for your records. Submit completed form via or fax completed form to Biological Laboratories, Inc. Attn: Antigen Receiving Dept. 255 Industrial Blvd. Everett, PA Lampire Project Management Approval Lampire Technical Support Approval SP200 38F1 Rev 2 Page 2 of 2 BIOLOGICAL LABORATORIES Tel: Fax: lampire@lampire.com

22 Hybridoma Project Initiation Form 1. General Information Company / Institution: Investigator Name: Contact Name: Phone: Fax: Project Name / Number: PO# Credit Card# (Name on Card & Exp ): 2. Billing / Shipping Information Accounts Payable contact: Company: Dept./Bldg./Room#: Address: Ship to contact: Company: Dept./Bldg./Room#: Address: City, State, Zip City, State, Zip Phone: Carrier (FedEx, UPS, etc.) Fax: Your carrier #: 3. Scope of Project What is the proposed end use for the antibody? Are there any special project requirements? 4. Antigen Information Immunogen name: Screening antigen name: Format (liquid, powder, etc.) Format (liquid, powder, etc.) Peptide Recombinant Peptide Type of Immunogen: Type of Antigen: Protein Other: Recombinant Protein Other: If Peptide, Describe Free Reactive Group: Cysteine Carboxyl Amino Immunogen MW (kda): Antigen MW (kda): Tag or conjugate name: Tag or conjugate name: Immunogen Source: Antigen Source: Storage conditions: 4 C -20 C -70 C Storage conditions: 4 C -20 C -70 C No. of containers: No. of containers: Volume per container: Volume per container: Concentration / Dosage Conc: Dose: Concentration: Potentially Biohazardous?* Yes No* Unknown Potentially Biohazardous?* Yes No* Unknown Handling instructions (required if biohazardous): Handling instructions (required if biohazardous): If attenuated, please provide documentation * If attenuated, please provide documentation * Has the antigen sequence been checked to determine whether it is conserved in mice? Yes No NOTE: If the antigen is conserved in mice, an animal of a different species may be recommended for generation of the monoclonal antibody. If this information is not known, will examine the antigen structure and determine whether or not the antigen is conserved and among which species it is conserved. Has the antigen been used previously to generate an immune response in an animal? Yes No If so, what species was used to generate the antibody? Antigen Structure (Peptide Sequence/Hapten with Spacer): SP200 38F2 Rev 1 Page 1 of 4 BIOLOGICAL LABORATORIES Tel: Fax: lampire@lampire.com

23 Hybridoma Project Initiation Form 5. Immunization Protocol Strain of Rodent for Immunization: ( does not use SCID or Nude mice for hybridoma development projects.) Balb/C Female Mice (9-12 weeks) Sprague Dawley Rats (200g) Other Number of Rodents to Immunize: (Projects typically begin with 5 or 10 mice or 3 rats) Mice: 5 10 Other Rats: 3 Other Recommended Injection Concentration and Volume: Please choose from one of the Immunization Protocols below. If a modified protocol is to be used, please describe. Standard Immunization Protocol: Day 0 - Pre-Bleed and 1st Injection (Complete Freunds) Day nd Injection (Incomplete Freunds) 1 Day Test Bleed Day ELISA Screen Modified Immunization Protocol (describe below): Additional injections and bleeds can be performed as needed. Additional injections will be given 18 days after the last bleed and will be followed by a test bleed 10 days after the injection unless otherwise directed. Screening of Bleeds is performed by ELISA. If you would like ELISA performed, please describe the screening strategy: 2 Do you wish to screen the test bleeds by the Final Assay Format? Yes No If Yes, test bleeds will be shipped to client (Mice ~10µL & Rats ~50µL). NOTE: Mice will be maintained at for a period of 3 months. After that time and with no continuing activity other than husbandry, a charge of $1/box of mice per day will be charged for husbandry of the animals. 1 performs all screening by ELISA. The assay verifies whether or not the animal has developed an immune response against the injected material. 2 If possible, recommends that Test Bleeds (and/or supernatant) be evaluated in the Final Assay Format. If the Test Bleeds do not perform in the Final Assay Format, the antigen may need to be modified or additional injections may need to be performed. 6. Specifications for Fusion and Screening How will the fusion candidate be selected: Lampire data Client data Both Rodents are boosted IP 3 consecutive days prior to fusion with the same injection concentration recommended in Section 5. Other immunization strategies are available upon request. If the following selections are dependent upon fusion data, will contact you to discuss after release of the data. Number of Growth Wells to select for subcloning: 3 Other Screening of Growth Wells is performed by ELISA. Should samples (1mL supernatant) be shipped to you for another screening format? Yes No Do you want to screen the Growth Wells by the Final Assay Format? If yes, how many at a time? ( suggests this be done after 1X cloning or within 1 week of shipment.) Yes No Criteria for selection of Growth Wells for subcloning: After subcloning, 3mL of supernatant will be provided. will be in contact with you to discuss future needs such as larger scale production, cell banking and purification. SP200 38F2 Rev 1 Page 2 of 4 BIOLOGICAL LABORATORIES Tel: Fax: lampire@lampire.com