A 2D LC-MS/MS Strategy for Reliable Detection of 10-ppm Level Residual Host Cell Proteins in Therapeutic Antibodies
| Autor: | David A. Michels, Alice Zhang, John T. Stults, Reed J. Harris, Donald E. Walker, Joseph Carver, Feng Yang, Jeannine Schoenfelder, Delia Li, X. Christopher Yu |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Residual Sensitivity and Specificity 01 natural sciences Analytical Chemistry 03 medical and health sciences Cricetulus Tandem Mass Spectrometry Lc ms ms Escherichia coli Animals Humans Chromatography Reverse-Phase Chromatography biology Chemistry 010401 analytical chemistry Ms analysis Antibodies Monoclonal 0104 chemical sciences 030104 developmental biology Biopharmaceutical biology.protein Cattle Antibody Drug Contamination |
| Zdroj: | Analytical Chemistry. 90:13365-13372 |
| ISSN: | 1520-6882 0003-2700 |
| Popis: | Methodologies employing LC-MS/MS have been increasingly used for characterization and identification of residual host cell proteins (HCPs) in biopharmaceutical products to ensure their consistent product quality and safety for patients. To improve the sensitivity and reliability for HCP detection, we developed a high pH-low pH two-dimensional reversed phase LC-MS/MS approach in conjunction with offline fraction concatenation. Proof-of -concept was established using a model in which seven proteins spanning a size range of 29-78 kDa are spiked into a purified antibody product to simulate the presence of low-level HCPs. By incorporating a tandem column configuration and a shallow gradient through the second-dimension, all seven proteins were consistently identified at 10 ppm with 100% success rate following LC-MS/MS analysis of six concatenated fractions across multiple analysts, column lots and injection loads. Using the more complex Universal Proteomic Standard 1 (UPS-1) as an HCP model, positive identification was consistently achieved for 19 of the 22 proteins in 8-12 ppm range (10 ppm ±20%). For the first time, we demonstrate an effective LC-MS/MS strategy that not only has high sensitivity but also high reliability for HCP detection. The method performance has high impact on pharmaceutical company practices in using advanced LC-MS/MS technology to ensure product quality and patient safety. |
| Databáze: | OpenAIRE |
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