Affinity resin selection for efficient capture of bispecific antibodies as guided by domain composition

Abstract

Affinity chromatography is a cornerstone of bispecific antibody (bsAb) purification, with resin selection playing a critical role in developing downstream processes to ensure both process efficiency and product quality. Unlike monoclonal antibody purification, where Protein A chromatography is the gold standard for antibody capture, affinity chromatography in bsAb purification is often employed not only for capture but also for the removal of hard-to-eliminate product-related impurities. This study demonstrates that affinity resin selection can be effectively guided by analysing the domain composition of the target bsAb molecule and its potential impurities. Using faricimab, a CH1-CL CrossMab, as a model, Protein L — an affinity resin targeting the variable region of the light chain — was predicted to be the most effective affinity chromatography due to its different binding avidity towards faricimab from its major product-related impurities. Validation through screening four different types of affinity chromatography, each binding to distinct regions of faricimab, confirmed this prediction. Under optimized elution conditions, the purification process achieved ∼73 % purity from ∼30 % in the culture with ∼86 % monomeric yield, as well as decent removal of host cell proteins and host cell DNA.