INVESTIGATION OF PROTEIN-PATTERNS IN MAMMALIAN-CELLS AND CULTURE SUPERNATANTS BY MATRIX-ASSISTED-LASER-DESORPTION IONIZATION MASS-SPECTROMETRY/

The direct protein profiling of mammalian cells and bacteria has a gro wing influence in biotechnology as a high information bearing method f or characterization of cells and cell states. Monitoring of proteins e xcreted in culture media not only serves to produce data on product yi eld and quality but provides important information on cell viability a nd nutrient supply that forms the basis for future process and express ion optimization, Fast and simple MALDI mass spectrometry approaches w ere developed to efficiently characterize such complex biological syst ems. Several mammalian cell lines including CHO DXB11, CHOSSF3, and hy bridomas were investigated; the lysis process, the sample pretreatment , and the matrix preparation were optimized for MALDI conditions, Init ial experiments to observe the success of protein translation in gene expression experiments were performed. Using MALDI-compatible detergen ts, it was possible to extend the mass range detectable by MALDI mass spectrometry from the current range of 16000 to 75000 Da. In this mass range, the data are complementary (offering a better mass accuracy) t o those obtained by SDS-PAGE electrophoresis experiments. These new me thods were used to monitor;a large-scale cultivation of hybridoma cell s expressing an antibody of the IgG type. The increase in whole antibo dy and antibody light-chain protein, 8650 Da, and the decrease of insu lin were followed during the monitoring period. Quantitative measureme nts of the IgG level during the cultivation compared favorably with th ose obtained by affinity HPLC.