Analysis of host cell protein residues in bevacizumab by Nano-UPLC-MS/MS and ELISA
10.13200/j.cnki.cjb.004481
- VernacularTitle:纳升超高效液相色谱-质谱联用技术及酶联免疫吸附试验分析贝伐珠单抗中宿主细胞蛋白残留
- Author:
LU Jingjing
- Publication Type:Journal Article
- Keywords:
Bevacizumab;
Host cell protein(HCP);
Nano ultra-performance liquid chromatography-tandem mass spectrometry(Nano-UPLC-MS/MS);
Enzyme-linked immunosorbent assay(ELISA)
- From:
Chinese Journal of Biologicals
2025;38(05):588-594+604
- CountryChina
- Language:Chinese
-
Abstract:
Objective To analyze the host cell protein(HCP) residues in bevacizumab injection and its downstream productsby nano ultra-performance liquid chromatography-tandem mass spectrometry(Nano-UPLC-MS/MS) and enzyme-linked immu-nosorbent assay(ELISA).MethodsThe samples were directly treated with trypsin, and the undigested antibodies wereprecipitated by heat treatment, which were then detected by Nano-UPLC connected with Q-Exactive Mass Spectrometer. Thecollected peptide information was compared with the total protein library of Chinese hamsters by MaxQuant software, and theHCP types were identified. The absorbance values of samples were detected by double-antibody sandwich ELISA, and theHCP content was calculated according to the four-parameter curve.ResultsAfter purification by downstream process, thelevels and types of HCPs decreased significantly. The content of HCPs in injection was 7 ppm, including 20 types, amongwhich 5 types of persistent residual HCPs were detected, and 4 types of high-risk HCPs were not detected. Compared withthe original drug, the HCP residues of the two drugs were both less than or equal to 15 ppm, and there were differences inHCP types due to different purification processes.ConclusionThe combination of Nano-UPLC-MS/MS and ELISA providesa comprehensive analytical strategy for the characterization and quantification of HCP, especially the types and distributionof persistent residual HCP and high-risk HCP, which can provide a reference for the process optimization, comparabilityresearch and safety risk assessment of products.
