Heath C, Dilwith R, Belfort G
Department of Chemical Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180-3590.
J Biotechnol. 1990 Jul;15(1-2):71-89. doi: 10.1016/0168-1656(90)90052-d.
The growth and antibody production of the SP2/0-derived hybridoma HB124 (ATCC) grown in media containing varying amounts of fetal bovine serum (FBS) were monitored using biochemical and flow cytometric methods. Hybridomas grown in 100 ml spinner flasks with RPMI-1640 containing varying amounts of serum demonstrated that cell growth, viability and IgG production show significant changes when serum content is decreased from 10.0 to 5.5 to 1.0 and 0.5%. A longer lag phase resulted when the lower serum content media were used. Cellular rates of glucose uptake showed a significant increase as serum levels were lowered. Similarly, exponential phase IgG production rates increased as the amount of serum was decreased, probably as a result of the decreased rate of exponential growth. Flow cytometric analysis showed a similar increase in cellular IgG content as medium serum levels declined. In contrast, the maximum IgG concentrations were found in flasks containing 1% FBS or above with the lowest concentration in the 0.5% FBS flask being due to the lower numbers of viable cells. Cells grown in microporous hollow fiber reactors were fed with medium containing serum which was decreased stepwise with time. Decreasing medium serum content stepwise from 10 to 2.5% resulted in increased antibody production. However, complete removal of serum from the medium resulted in a significant drop in antibody productivity. Cumulative antibody production was equivalent for cells grown entirely in medium containing 10% FBS and for those which experienced a drop to 2.5% FBS. To compare a defined serum-free medium preparation with medium containing 10% FBS, cells were again grown in batch suspension culture and analyzed. The growth rates were similar but there was a significant difference in IgG production rates. The serum-free culture exhibited both higher cellular production rates and higher IgG concentrations. These results indicate that decreasing medium serum content can adversely affect antibody yield because of lower cell viabilities, not because of lower production rates. Use of a defined serum-free medium, as done in this study, results in higher yields because of a higher IgG production rate as well as good cell growth and viability.
采用生化和流式细胞术方法，监测了在含有不同量胎牛血清（FBS）的培养基中生长的源自SP2/0的杂交瘤HB124（美国典型培养物保藏中心）的生长和抗体产生情况。在含有不同量血清的RPMI-1640培养基中，于100 ml旋转瓶中培养杂交瘤，结果表明，当血清含量从10.0%降至5.5%、1.0%和0.5%时，细胞生长、活力和IgG产生均出现显著变化。使用较低血清含量的培养基时，延迟期更长。随着血清水平降低，细胞葡萄糖摄取率显著增加。同样，指数期IgG产生率随着血清量的减少而增加，这可能是指数生长速率降低的结果。流式细胞术分析表明，随着培养基血清水平下降，细胞内IgG含量有类似增加。相比之下，在含有1% FBS或更高浓度FBS的培养瓶中发现最大IgG浓度，而在0.5% FBS培养瓶中浓度最低，这是由于活细胞数量较少。在微孔中空纤维反应器中培养的细胞，用含血清的培养基喂养，血清含量随时间逐步降低。将培养基血清含量从10%逐步降至2.5%，导致抗体产生增加。然而，培养基中完全去除血清会导致抗体生产力显著下降。完全在含10% FBS的培养基中生长的细胞与经历血清降至2.5%的细胞的累积抗体产量相当。为了将一种明确的无血清培养基制剂与含10% FBS的培养基进行比较，细胞再次在分批悬浮培养中生长并进行分析。生长速率相似，但IgG产生率有显著差异。无血清培养显示出更高的细胞产生率和更高的IgG浓度。这些结果表明，降低培养基血清含量可能会因细胞活力降低而对抗体产量产生不利影响，而非因为产生率降低。如本研究中所做的那样，使用明确的无血清培养基，由于更高的IgG产生率以及良好的细胞生长和活力，可导致更高的产量。
