Biologically manufactured monoclonal antibodies (mAb) can strongly vary in their efficacy and affinity. Therefore, engineering and production of the mAb is highly regulated and requires product monitoring, especially in terms of N-glycosylation patterns. In this work, we present a high-throughput matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) method based on a microarray technology to monitor N-glycopeptides of IgG1 produced in a perfusion cell culture. A bottom-up approach combined with zwitterionic-hydrophilic interaction liquid chromatography for sample purification was used to determine the day-by-day variation of the terminal galactose within two major N-glycoforms. Our results show that microarrays for mass spectrometry (MAMS) are a robust platform for the rapid determination of the carbohydrate distribution. The spectral repeatability is characterized by a low coefficient of variations (1.7% and 7.1% for the FA2 and FA2G1 structures, respectively) and allows to detect the N-glycosylation variability resulting from operating conditions during the bioreactor process. The observed trend of released N-glycans was confirmed using capillary gel electrophoresis with laser-induced fluorescence detection. Therefore, the microarray technology is a promising analytical tool for glycosylation control during the production process of recombinant proteins.
Keywords: MALDI-MS; Microarray for mass spectrometry (MAMS); Monoclonal antibody; N-linked glycosylation; Perfusion bioreactor.
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