Mycoplasma (-) Negative Testing
Cell lines received should have documentation linking the frozen vials to mycoplasma (-) negative test results. If such documentation is not available, the cell line will be quarantined. Quarantined cell lines will be thawed in a dedicated cleanroom suite and tested extensively for microbes such as bacteria, fungi, mycoplasma and other adventitious agents. The creation of a mycoplasma (-) negative certified working cell stock (CWCS) enables the cell line to be thawed in the manufacturing cell culture suites. The creation of certified working cell stock does not assure that the cell line is optimized for In vitro production of monoclonal antibodies or proteins in bioreactors.
Clonality and Productivity Testing
To determine if a cell line is optimal for In vitro production of monoclonal antibodies, a clonality and specific productivity rate (SPR) assay should be performed to determine the percentage of cells in the culture producing antibodies of a similar titer. To maximize monoclonal antibody production, a cell line optimization procedure should be performed during the creation of a master cell bank (MCB). If a cell line optimization procedure is not performed, non-producing cells may be selected over antibody producers in the bioreactor or during adaptation to protein free, animal origin free media.
Cell Line Optimization
Using the limiting dilution cloning technique, parental cell cultures are cloned and tested for antibody production. After each round of limiting dilution cloning, cells are tested for antibody production and the results are analyzed using a statistical method to determine when the cell line is optimized for In vitro production of monoclonal antibodies.
Mammalian cell lines including CHO, NSO, HEK-293 and hybridomas may be adapted for cell culture in chemically defined, protein free, animal origin free medium. Bioreactor production of recombinant proteins and antibodies is routinely carried out in a protein free environment. This often facilitates downstream processing procedures assuring that the final product is manufactured in an animal origin free system.