Antibody Isolation from Human Synthetic Libraries of Single-Chain Antibodies and Analysis Using NGS

Abstract

Antibody libraries came into existence 30 years ago when the accumulating sequence data of immunoglobulin genes and the advent of PCR technology made it possible to clone antibody gene repertoires. Phage display (most common) and additional display and screening technologies were applied to pan out desired binding specificities from antibody libraries. As other antibody discovery tools, phage display is not an off-the-shelf technology and not offered as a kit but rather requires experience and expertise for making it indeed very useful.Next-generation sequencing (NGS) coupled with bioinformatics is a powerful tool for analyzing large amount of DNA sequence output of the panning. Here, we demonstrate how NGS analysis of phage biopanning (phage-Seq) of complex antibody libraries can facilitate the antibody discovery process and provide insights regarding the biopanning process (see Fig. 1).

Keywords: CDRs (complementarity determining regions); FR (variable framework region); HTS (high-throughput sequencing); NGS (next-generation sequencing; Phage display; Phage-Seq; Single-chain antibodies; Synthetic library; VH (variable region of antibody heavy chain); VL (variable region of antibody light chain); scFv (single-chain variable fragment).