Clonal Variant Analysis of Antibody Engineering Libraries

In vitro antibody evolution is a powerful technique for improving monoclonal antibodies. This can be achieved by generating artificial diversity on an antibody template, which can be done using different in vitro diversification techniques. The resulting libraries consist of single- or multimutant variants of a defined antibody template that are screened for improved function using antibody display. Here, we describe a bioinformatic protocol for tracking synthetic antibody variants using high-throughput sequencing across screening rounds, enabling efficient high-throughput interpretation of the function of individual mutations in sorted antibody display libraries. The protocol enables a user to achieve precision analysis and interpretation of clonal antibody variant data for discovery purposes, especially for high-throughput antibody engineering or optimization against target antigens.