Primer design and inverse PCR on yeast display antibody selection outputs

Abstract

The display of antibodies on the surface of Saccharomyces cerevisiae cells enables the high-throughput and precise selection of specific binders for the target antigen. The recent implementation of next-generation sequencing (NGS) to antibody display screening provides a complete picture of the entire selected polyclonal population. As such, NGS overcomes the limitations of random clones screening, but it comes with two main limitations: (1) depending upon the platform, the sequencing is usually restricted to the variable heavy chain domain complementary determining region 3 (HCDR3), or VH gene, and does not provide additional information on the rest of the antibody gene, including the VL; and (2) the sequence-identified clones are not physically available for validation. Here, we describe a rapid and effective protocol based on an inverse-PCR method to recover specific antibody clones based on their HCDR3 sequence from a yeast display selection output.