Ligand Selection by Combination of Recombinant and Cell Panning Selection Techniques.

Abstract

High-throughput protein selection methods are a cornerstone for protein engineering and pharmaceutical development. Traditional high-throughput selection strategies rely largely on recombinant antigen to generate target specificity. Though effective, this selection strategy can be limited by soluble target quality, particularly in the case of recombinant extracellular domains of transmembrane proteins. Recent advances in cell-based selection techniques provide new opportunities for improving the outcomes of ligand selection campaigns but can introduce technical challenges in maintaining antigen specificity due to the heterogeneity of biomacromolecule expression on the mammalian cell surface. Here, we describe a combination technique using recombinant antigen to "train" library target specificity followed by cell panning selections to ensure that isolated ligands bind cell-expressed target, as well as a facile microscopy technique for assessing target specificity on a clonal basis without the need to produce soluble ligand.