Mouse scFv Library Generation and Production
AbGenics provides specialized mouse short chain variable fragment (scFv) antibody generation services against specific antigens provided by the clients. Antibodies libraries are generated from immunized mice and monoclonal anitbody fragments are isolated by phage display, screened and assayed for specific biological activity. The scFvs are purified by affinity chromatography and provided to end users in milligram quantities. The mouse antibody development takes place through the following steps:
- Immunisation of the mice : Antigen required, 2 mgs. 5-6 animals are immunised subcutaneously or intradermally with the antigen in adjuvant.
- Booster doses and final bleeding: Booster doses are given at 14, 28, 42 days from the first immunization. Antibody titres are checked prior to the final bleed, mice are sacrificed and spleens harvested and processed.
- Generation of the library : The cDNA is isolated from the spleens the light and the heavy chains are extracted out by a PCR reaction and brought together by overlap-extension PCR to generate scFv and a library is prepared in E coli.
- Panning and screening: The library is panned and screened by phage display to generate monoclonal scFvs against specific targets.
- Production and purification of the antibodies : The antibodies are produced in small amounts in bacterial/yeast systems, purified by affinity chromatography and supplied to the end users in mg quantities.