Cell Banking, Optimization and Adaptation

The first step in a quality manufacturing system is to assure creation of master and working cell banks. Leinco Technologies uses a tiered cell banking system to ensure a consistent supply of characterized cells for use in continuous cycle manufacturing campaigns. Cell banking work is performed in dedicated cleanroom suites using standard operating procedures and data from the preparation of cell banks is recorded in detailed batch records.

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. Creation of the 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.

Adaptation

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.

Master and Working Cell Banks

The master cell bank (MCB) is created from a single cell source optimized for monoclonal antibody production. Cells are expanded to a level required to freeze an appropriate number of vials based on demand for their use. Master cell banks are tested for recovery viability after cryopreservation. A master cell bank is biosafety tested and the cell line is authenticated. The scope of biosafety testing and authentication is determined based on the client-specific application and advice from the worldwide regulatory bodies.

The working cell bank (WCB) is created from the expansion of a master cell bank vial. Expansion and cryopreservation of the working cell bank depends on the demand for use. It is the working cell bank that is used by cell culture manufacturing scientists to produce the seed culture for inoculation of bioreactors. Leinco Technologies offers custom In vitro Antibody and Protein Production in stirred tank and hollow fiber bioreactors.

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