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Several analytical techniques were applied to obtain information about the stability of expression, yield and integrity of monoclonal antibodies (Mabs) produced by hybridoma cells in homogeneous continuous perfusion culture systems. An isotype-specific spot enzyme-linked immunosorbent assay (spot-ELISA) was developed to determine the amount of antibody-producing cells and isotype switch variants. A flow cytometric (FC) procedure was applied to determine the relative cytoplasmic IgG content. Further, ELISA and isoelectric focusing (IEF) were applied for the analysis of the Mab secreted in the culture fluid. Spot-ELISA, FC analysis, ELISA and IEF were used to monitor the antibody formation of the RIV6 hybridoma in a homogeneous continuous culture system. This cell line turned out to be unstable in this culture system with respect to cell properties and product formation. |
