Development of an alternative method for the identification and production of antigen-specific monoclonal antibodies (65.16)

Abstract

Abstract A critical step in the production of monoclonal antibodies (Abs) is the initial identification of the antigen-specific Abs, which is usually performed by multiple rounds of panning in both hybridoma and phage display. We have developed an alternative method that allows for the rapid and direct identification of antigen-specific Abs from peripheral blood via high-throughput immune repertoire sequencing and LC MS/MS peptide matching. Our B-cell repertoire technology combines novel amplicon rescued multiplex PCR (arm-PCR, patent pending) with high-throughput gene sequencing to access the sequence of a broad spectrum of heavy and light chain V-regions. As a proof of concept, we have sequenced the immune repertoire of 2 healthy individuals at various time points after administration of the 2009-2010 seasonal influenza vaccine. Antigen-specific Abs were purified directly from immune peripheral blood serum and identified using LC MS/MS peptide sequencing, exploiting the B-cell repertoire gene sequencing results as a database for identification. During our study, several unique peptides were successfully matched for each individual’s response to both Flu A strains in the vaccine. Future work includes the development of a method to rapidly clone and express these identified Abs in a human in vitro glycoexpression system. The recombinant Abs will then be tested for their ability to bind flu hemagglutinin, demonstrating the utility of our technology towards the production of mAbs.