哺乳动物细胞培养
哺乳动物细胞培养是一种面向研究、临床和制药应用的重要工具,可将从动物组织分离得到的细胞培养扩增,用于细胞生物学和疾病研究以及抗体、蛋白质和疫苗生产。这些永生化的哺乳动物细胞系可在体外长期生长,常用作复杂生物学的简化模型。
哺乳动物细胞的体外生长
细胞生长需要复杂的营养混合物,包括糖类、氨基酸、白蛋白、维生素、矿物质和生长因子等。细胞培养基是哺乳动物细胞体外培养所需营养物质的主要来源。而培养箱则为细胞培养提供无菌环境,保持37 °C的理想温度以及5%的 CO2浓度,使箱内pH水平接近哺乳动物血液。
除了某些类型的血细胞悬浮培养外,多数哺乳动物组织的细胞需要贴壁培养才能正常地分裂和生长。经处理过的聚苯乙烯或玻璃材质培养瓶、培养板和过滤器可为细胞贴壁培养提供生长表面。而多聚D赖氨酸等合成聚合物及胶原蛋白等细胞外基质,则可为细胞粘附提供骨架以促进贴壁。
哺乳动物细胞培养技巧
无菌技术可保护细胞体外生长免受污染。为确保细胞健康并增殖,哺乳动物细胞培养需要进行传代培养,在细胞达到一定汇合度后稀释并更换耗尽的培养基。细胞贴壁培养时,将细胞转移到新培养瓶或培养板重贴壁之前,必须通过胰蛋白酶化等酶解法或刮铲等机械法先将细胞解离。然后采用血细胞计数板和自动细胞计数仪进行细胞计数来测定细胞活力并确定传代接种密度和适合下游过程的细胞密度。培养细胞可用液氮冻存生成种子库,需要时解冻即可制备。
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