Antibody Batch Cloning.

Abstract

The antigen-binding ability of each antibody clone selected by phage display is usually initially ranked by a screening ELISA using monovalent scFv antibody fragments. Further characterization often requires bivalent antibody molecules such as IgG or scFv-Fc fusions. To produce these, the V region encoding genes of selected hits have to be cloned into a mammalian expression vector and analyzed as a bivalent molecule, requiring a laborious cloning procedure. We established a high-throughput procedure allowing rapid screening of candidates in bivalent formats. This protocol allows for the parallelized cloning of all selected antibody fragments into a mammalian expression vector in the 96-well plate format. The bivalent antibody molecules can then be produced and purified in 96-well plates for further analysis in microtiter plate assays.