Ultrasensitive method for profiling host cell proteins by coupling limited digestion to ProteoMiner technology.

Host cell proteins (HCPs) are process-related impurities that remain in therapeutic protein -at trace levels. HCPs must be closely monitored because they may be detrimental to drug product quality. Liquid chromatography coupled to mass spectrometry (LC-MS) is a powerful tool for detecting individual HCPs; however, HCP-derived peptides can be four to ten orders of magnitude less abundant than therapeutic protein-derived peptides in drug products, thus posing a major challenge in LC-MS detection. We previously demonstrated that low abundant HCPs can be enriched several hundreds fold through ProteoMiner. This study further improved the degree of enrichment by coupling limited digestion to ProteoMiner technology (PMLD). HCPs with low abundance were enriched 7694-fold, thus enabling detection of HCPs at concentrations as low as 0.002 ppm. A total of 850 HCPs were detected with high confidence from a NIST monoclonal antibody preparation, a number 40% higher than previously reported.