Protocols
Phase I: Immunization
- You provide antigen or identify an antigen source. The probability of identifying useful antibodies improves with increasing antigen purity. We can assist in the preparation and purification of antigens.
- Haptens, peptide and weak or non-immunogenic antigens may require conjugation to carrier proteins. We offer conjugation to carrier molecules such as KLH, BSA and ovalbumin and can conjugate to the carrier molecule of your choice.
- Mice are immunized over three months. Sera is periodically examined for specific antibodies. Your end use of the antibody determines the most appropriate assay used, but can include ELISA, Western blot, RIA or other specified assay formats. Sera is available for further testing.
Alternative immunization protocols can be explored if serum titers are determined to be unacceptable prior to initiating fusion procedures.
Timing: 10-12 weeks, shorter if sera titers are acceptable.
Pricing: Determined by the number and complexity of assays utilized.
Phase II: Cell Fusion and Screening
- Splenocytes from mice with acceptable sera antibody activity are fused with a myeloma cell line.
- Resulting hybridomas are screened with the same assays used to demonstrate serum antibody performance in Phase I.
- Hybridoma culture supernatant is available to the client for further testing.
- Positive hybridomas are frozen and cryopreserved.
Timing: Typically 5-6 weeks if a single cell fusion yields useful antibodies. Dependent on the number of antibodies needed and antibody sensitivity and specificity requirements.
Pricing: Determined by the number of antibodies required and the number and complexity of screening assays.
Phase III: Cloning, Antibody Production and Purification
- Limiting dilution cloning of the hybridoma cell line is performed to insure monoclonality of the cell line.
- Clones are tested using the same assays used in Phases I and II.
- Clones are frozen and cryopreserved.
- Clones are injected into mice for production of antibody in ascites.
- Antibody is purified from ascites and supplied at a concentration and in a buffer specified by the client.
- The isoelectric point and the purity of the antibody is determined. A Certificate of Analysis is supplied for each purified antibody.
- Resulting antibodies and hybridoma cell lines are the property of the client. Storage and shipment of these materials is directed by the client.
Timing: Approximately 10-12 weeks.
Pricing: Dependent on number of hybridomas cloned and number of antibodies produced in ascites and purified.
Standard 6 to 9 month program estimated
$7500 to $15000