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Lasunskaia, E.B. et al.

Lasunskaia, Fridlianskaia, Darieva, da Silva, Kanashiro, Margulis,

Transfection of NS0 myeloma fusion partner cells with HSP70 gene results in higher hybridoma yield by improving cellular resistance to apoptosis.

Mouse myeloma NS0 cells widely used in hybridoma technology lack the expression of a major stress protein Hsp70 which is the principal component of the basic cellular defense mechanism. These cells rapidly undergo apoptosis at the late-stationary phase of batch culture following nutrient exhaustion. Since Hsp70 was recently demonstrated to protect cells against numerous apoptotic stimuli, the aim of the present study was to examine the protective potential of the protein expression in engineered myeloma NS0 cells and in resulting hybridomas. Myeloma cells were transfected with the hsp70 gene under beta-actin gene promoter. To imitate harmful conditions that hybridoma or myeloma cells often experience when cultivated in large scale for an antibody production, NS0(wt) and NS0(hsp70) cell cultures were maintained without changing the medium for a few days, and the expression of apoptotic markers has been studied. It was found that long-term cultivation induced apoptosis in original cells manifested by typical nuclei fragmentation, DNA ladders and activation of caspase-3. In contrast, in transfected cells under the same conditions the outcome of apoptosis was postponed for 24 hours. Most relevant was that the fusion of transfected myeloma cells with immune splenocytes resulted in twofold hybridomas output compared with wild-type fusion partner. Almost half of the hybridomas continued to be hsp70-positive and maintained higher robustness in culture. The level of monoclonal antibodies production by hybridoma cells obtained with the use of NS0(wt) and NS0(hsp70) was similar, however, the secreted product was better preserved in culture supernatants of Hsp70-positive cells. It is concluded that transfection of mouse myeloma cells with the hsp70 gene can be a novel means to increase hybridoma yield and reduce the sensitivity of myeloma and hybridoma cells to culture conditions insults accompanying monoclonal antibody production.[1]

References

Transfection of NS0 myeloma fusion partner cells with HSP70 gene results in higher hybridoma yield by improving cellular resistance to apoptosis. Lasunskaia, E.B., Fridlianskaia, I.I., Darieva, Z.A., da Silva, M.S., Kanashiro, M.M., Margulis, B.A. Biotechnol. Bioeng. (2003) [Pubmed]

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