Abstract��
OBJECTIVE To optimize the culture temperature of recombinant CHO-DG44 cells expressing tumor necrosis factor receptor-Fc fusion protein and to determine the bioactivity of expressed protein at the best culture temperature. METHODS Recombinant CHO cells were batchly cultured at three different temperatures (37 �棬37 �� shifting to 31 �� and 31 ��). Samples were daily tested for cell densities�� viabilities�� glucose concentration�� lactic acid concentration and TNFR-Fc fusion protein concentration�� then the best culture temperature was chosen to scale up the fermentation volume��10�� 50�� 200 mL�� and 2 L��. TNFR-Fc fusion protein was purified with Protein A affinity chromatography��determined for relative molecular mass��purity and neutralizing activity by SDS-PAGE��SE-HPLC and WST-8 separately. RESULTS The culture condition of 37 �� shifting to 31 �� resulted in the maximum viable cell density��high viability and long culture time of the cells as well as high TNFR-Fc protein productivity��and this culture procedure could be successfully applied to the scale-up of recombinant CHO cells. The purified fusion protein��with a relative molecular mass about 150 000�� reached the purity of more than 93% and neutralized the cytotoxic effect of TNF��. CONCLUSION Compared to the traditional culture temperature of 37 �棬the culture temperature of 37 �� shifting to 31 �� can obviously improve the TNFR-Fc fusion protein output by recombinant CHO-DG44 cells. And this technique can be applied to scale-up�� which will lay a foundation of large scale industrial production.
LEI Yun,
XIE Kui,
HONG Gang etc
.Optimization of the Culture Temperature of Recombinant CHO-DG44 Cells Expressing Tumor Necrosis Factor Receptor-Fc[J] Chinese Pharmaceutical Journal, 2013,V48(2): 90-95
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2013, Vol. 48 
