(242c) Rapid Generation of Monoclonal Antibodies without Screening by Exploiting High-Throughput DNA Sequencing of Immunized Repertoires

Isolation of antigen-specific monoclonal antibodies (MAbs) is reliant on screening-based approaches such as B cell immortalization and surface display of recombinant antibody libraries. Here we describe a method for direct isolation of (MAbs), which circumvents the need for screening by exploiting the highly enriched, antigen-specific antibody repertoire of plasma cells - the terminal state of B cell differentiation. We utilized high-throughput DNA sequencing of antibody variable light and heavy regions (VL, VH) derived from mRNA transcripts of the plasma cell population present in the bone marrow of immunized mice. We found that the VL and VH gene repertoire was highly polarized. The most abundant and unique VL and VH genes were paired according to their relative abundance within the repertoire and rapidly and efficiently synthesized via an oligonucleotide and PCR assembly process under the control of automated liquid handling robots. Following recombinant expression in bacterial and mammalian systems, it was confirmed that antibodies were overwhelmingly (>75%) antigen specific, in some cases reaching nanomolar and sub-nanomolar affinities. We show that this simple approach enables the rapid generation of panels of high quality MAbs to a variety of antigens.