Determining Residual Host Cell Antigen Levels in Purified Recombinant Proteins by Slot Blot and Scanning Laser Densitometry

Abstract

The production of recombinant proteins for therapeutic and vaccine uses necessitates the analysis of purified products for residual host cell antigens. Various assays to quantify these antigens in both in-process streams and purified bulk solutions have been described (1–3). Process-specific assays utilize those host cell antigens most likely to copurify with the protein of interest, whereas generic assays use all recoverable host cell antigens, thereby testing for the presence of any antigen during protein production and that might therefore be recovered with the purified protein. With limited utility of commercially available assay kits, we developed a fast, simple, quantitative, and highly sensitive method based on immunoblotting to accurately determine residual host cell antigen levels. Samples are applied directly to a membrane, and signals are measured by densitometry. This assay results in very low-background signals, providing accuracy and precision at the low nanogram/milliliter level, comparable to reported enzyme-linked immunosorbent assay (ELISA)-based methods.