Sensitive detection and quantification of residual host cell proteins (HCPs) represent a critical step in the design of robust, well-controlled manufacturing processes that yield high-quality biopharmaceuticals. LC-MS-based methods are becoming a routine approach for HCP analysis where residual HCPs can be detected, identified, and quantified directly.
In this video, we present the results obtained in a comparison study using a generic, highly sensitive LC-MS-based assay for identification and quantification of HCPs in two highly purified monoclonal antibodies, an innovator Infliximab, Remicade®, and its biosimilar version, Inflectra™.
The HCP assay used for this study relies on: 1) proteolytic digestion; 2) two-dimensional chromatographic separation (high pH/low pH) by RP/RP UPLC; 3) high-resolution ESI-mass spectrometric detection coupled with ion-mobility separation (IMS) of peptide precursors; 4) drift-time specific fragmentation of peptide precursors using a fixed collision energy; and 5) database searching for HCP identification and quantification.
Two common HCPs are identified in both innovator and biosimilar Infliximab samples with concentrations ranging from 10 to 100 ppm. Our results show that the HCP levels of the biosimilar, Inflectra, are 2–4-fold higher than the innovator Infliximab. These results indicate that LC-MS assays are now able to attain comparable sensitivity to traditional HCP assays (e.g. ELISA), while offering the unique advantage in providing unambiguous HCP identification.