Ozturk S S, Palsson B O
Department of Chemical Engineering, University of Michigan, Ann Arbor 48109.
J Biotechnol. 1990 Nov;16(3-4):259-78. doi: 10.1016/0168-1656(90)90041-9.
A murine hybridoma cell line (167.4G5.3) was cultivated in batch mode with varying inoculum cell densities using IMDM media of varying fetal bovine serum concentrations. It was observed that maximum cell concentrations as well as the amount of monoclonal antibody attainable in batch mode were dependent on the inoculum size. Specifically, cultures with lower inoculum size resulted in lower cell yield and lower antibody concentrations. However, in the range of 10(2) to 10(5) cells per ml, the initial cell density affected the initial growth rate by a factor of only 20%. Furthermore, specific monoclonal antibody production rates were independent of initial cell density and the serum concentration. Glutamine was the limiting nutrient for all the cultures, determining the extent of growth and the amount of antibody produced. Serum was essential for cell growth and cultures with initial cell concentrations up to 10(6) cells per ml could not grow without serum. However, when adapted, the cells could grow in a custom-made serum-free medium containing insulin, transferrin, ethanolamine, and selenium (ITES) supplements. The cells adapted to the ITES medium could grow with an initial growth rate slightly higher than in 1.25% serum and the growth rate showed an initial density dependency-inocula at 10(3) cells per ml grew 30% slower than those at 10(4) or 10(5). This difference in growth rate was decreased to 10% with the addition of conditioned ITES medium. The addition of conditioned media, however, did not improve the cell growth for serum-containing batches.
使用不同胎牛血清浓度的IMDM培养基，以分批模式培养鼠杂交瘤细胞系（167.4G5.3），接种细胞密度不同。观察到分批模式下可达到的最大细胞浓度以及单克隆抗体的产量取决于接种量。具体而言，接种量较低的培养物导致细胞产量较低和抗体浓度较低。然而，在每毫升10²至10⁵个细胞的范围内，初始细胞密度对初始生长速率的影响仅为20%。此外，特异性单克隆抗体的产生速率与初始细胞密度和血清浓度无关。谷氨酰胺是所有培养物的限制性营养物质，决定了生长程度和产生的抗体量。血清对细胞生长至关重要，初始细胞浓度高达每毫升10⁶个细胞的培养物在无血清条件下无法生长。然而，经过适应后，细胞可以在含有胰岛素、转铁蛋白、乙醇胺和硒（ITES）补充剂的定制无血清培养基中生长。适应ITES培养基的细胞可以以略高于1.25%血清中的初始生长速率生长，并且生长速率显示出初始密度依赖性——每毫升接种10³个细胞的接种物比接种10⁴或10⁵个细胞的接种物生长慢30%。添加条件ITES培养基后，这种生长速率差异降至10%。然而，添加条件培养基并没有改善含血清批次的细胞生长。
