![]() ![]() There is an urgent need for a discovery engine that adequately mines the natural B cell diversity to rapidly isolate antigen-specific antibodies from human patients. Both of these methods therefore suffer from low screening throughput that overwhelmingly under-samples the ~10 7 B cells obtained from a typical blood draw. Moreover, validating leads requires gene synthesis, cloning, and expression which can create a severe bottleneck in the number of candidates that can be functionally assessed 14. Though the number of cells that can be sequenced with this method is high, inferring antigen-specificity from sequencing information is very challenging, especially from humans who are constantly exposed to a large diversity of antigens. A more recent development uses next-generation sequencing (NGS) to profile the paired repertoire from millions of B cells 12, 13. However, maintaining large numbers of monoclonal cultures for extended periods of time is laborious, expensive and limits the screening to a fraction of the B cell repertoire 11. First, B cells can be cultured in individual wells and their conditioned media screened for function, or their antibody genes directly cloned. These methods generally fall into two categories. Technologies that preserve this native pairing are therefore best suited to recapitulate the functional characteristics of naturally produced antibodies. Adding to this challenge is the fact that antibodies are heterodimeric proteins whose specificities are encoded by unique pairs of heavy-chain and light-chain transcripts. However, the B cells producing these therapeutic antibodies tend to be rare in convalescent patients, making their discovery very challenging. Because these antibodies are elicited in human responses to disease, they are believed to be safer, less immunogenic and in general more translatable to human therapy 10. Whereas most approved therapeutic antibodies have been obtained by engineering rodent antibodies, highly potent antibodies have recently been identified within humans against many diseases, including microbial infection 2, 3, 4, 5, autoimmunity 6, 7, 8, and cancer 9. Most isolated antibody sequences were not detected by next-generation sequencing of the paired repertoire, illustrating how this method can isolate extremely rare leads not likely found by existing technologies.Īntibodies are among the fastest growing therapeutic classes within the biopharmaceutical industry 1. Within 4 weeks we progressed from B cell isolation to a panel of unique monoclonal antibodies, including seven that displayed broad reactivity to different clinically relevant influenza hemagglutinin subtypes. We used this approach to construct natively paired phage-display libraries from healthy donors and drove selection towards cross-reactive antibodies targeting influenza hemagglutinin. Here we show the encapsulation of two million primary B cells into picoliter-sized droplets, where their cognate V genes are fused in-frame to form a library of scFv cassettes. However, methods for mining primary antibody-expressing B cells are limited in their ability to rapidly isolate rare and antigen-specific binders. The human antibody repertoire is increasingly being recognized as a valuable source of therapeutic grade antibodies. ![]()
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