For information about our custom polyclonal antibody service, click here.

Custom Monoclonal Antibody Service

Custom monoclonal antibody production at Allied normally takes 4-6 months and is divided into 3 phases. The client has the option of halting the hybridoma production prior to the beginning of the next phase. Allied Biotech will not proceed to the next phase of the project without written (e-mail) confirmation from the client. The client is invoiced for each phase at its initiation. The amount of antigen needed for immunizations of 5 mice is 1mg to 1.5mg and an additional 0.8mg to 1mg is needed for ELISA screening.

Hybridoma Development

Step 1, Antigen Preparation: the antigen used for immunization can be the following forms: purified proteins (500ug-2mg) peptides (5-10mg). We can conjugate your peptide or small protein to a carrier protein such as BSA or KLH.
Step 2, Immunization: In general, 5 Balb/c mice will be used. We begin with a 50 microgram priming dose followed by several boosting doses of 50 microgram each. This regime will continue until a sufficient antiserum titer is achieved.
Step 3, Serum Screening: Test bleeds are routinely performed and evaluated by direct binding ELISA with your antigen. Optimal ELISA conditions are developed to ensure the proper evaluation of future clones. In addition, test bleeds can be sent to your lab for evaluation in other assays.
Step 4, Evaluation: Data is evaluated jointly with Rockland staff to determine the performance of antiserum at this point. Antiserum titers and specific assay performance must be acceptable to continue the project.

Step 5, Fusion: One half of splenocytes from the mouse which responds the best to the immunizations as determined by ELISA or another assay will be fused with mouse myeloma cells. The remaining spleen cells will be frozen and kept in our facility.
Step 6, Screening: The fusion is plated into a stack of ten 96 well plates. Plates are monitored for growth and fed weekly. Wells with cell growth are screened in 2 to 3 weeks. Our standard screen method is ELISA. A small amount of purified protein is required for coating the EIA plates. If it’s not available, we shall an alternative, suitable method.

Step 7, Subcloning: Cells from 5 parental clones will be subcloned twice by limited diluting and screened by ELISA to establish stable hybridoma cell line.
Step 8, Expanding, freezing and growing of hybridoma cell lines: Up to 10 of the original different positive clones and 4 of the first subclones will be frozen 2 vials each. Final stable hybridoma cell lines will be frozen 10 vials each, and will be stored for up to six months.
Total Cost for Phase I through III (Steps 1 through 8): $6100
Time: Approximately 4-6 months depending on the response to the antigen.

Additional Services

Step 7 of Allied's complete service includes up to 5 positive parental clones. Additional subcloning and expansion can be performed at a rate of $195.00 per clone.

Precious clones can be maintained in liquid nitrogen in our monitored facility at a rate of $89.00 per clone per year.

The hybridoma cell lines developed by Rockland or from other sources can be tested for any Mycoplasma contamination at a rate of $150.00 per clone. 

Ascites Production

Ascites is produced in Balb/c strain mice. Optimized methods are used to generate tumors and accumulate ascitic fluid, including the quantity of cells and volume of inocula. These methods generate high yields of antibody that can be be further processed or characterized depending on your particular requirements.

Please inquire for larger quantities of mice for ascites production.

© 2003 Allied Biotech, Inc.