Exploiting next-generation sequencing in antibody selections - a simple PCR method to recover binders

• Published in: mAbs 2020-01, Vol.12 (1), p.1701792-1701792
• Main Authors: Ferrara, Fortunato, Teixeira, Andre A, Naranjo, Leslie, Erasmus, M Frank, D'Angelo, Sara, Bradbury, Andrew R M
• Format: Article
• Language: English
• Subjects: antibody; antibody libraries; Drug Discovery; Genetic Vectors; High-Throughput Nucleotide Sequencing; Humans; NGS (Next-generation sequencing); Peptide Library; phage display; Polymerase Chain Reaction - methods; Protein Binding; Recombinational DNA Repair; Single-Chain Antibodies - genetics; yeast display; Yeasts - genetics
• ISSN: 1942-0862; 1942-0870
• DOI: 10.1080/19420862.2019.1701792

Antibody discovery using display technologies such as phage and/or yeast display has become acornerstone in many research and development projects, including the creation of new drugs for clinical use. Traditionally, after the selection phase, random clones are isolated for binding validation and Sanger sequencing. More recently, next-generation sequencing (NGS) technology has allowed deeper insight into the antibody population after aselection campaign, enabling the identification of many more specific binders. However, this approach only provides the DNA sequences of potential binders, the properties of which need to be fully elucidated by obtaining corresponding clones and expressing them for further validation. Here we present arapid novel method to harvest potential clones identified by NGS that uses asimple PCR and yeast recombination approach. The protocol was tested in selections against three different targets and was able to recover clones at an abundance level that would be impractical to identify using traditional methods.