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Metabolic screening of mammalian cell cultures using well-plates

机译：使用孔板对哺乳动物细胞培养物进行代谢筛选

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摘要

For use in a broad spectrum of cell culture applications, we have devised a novel method, termed High-Throughput Metabolic Screening (HTMS), with which to more rapidly screen the overall activity of major metabolic pathways of mammalian cells. This current protocol uses adaptations of theoretical and experimental techniques from metabolic and cell culture engineering. First, HTMS males use of a simplified metabolic network for metabolic flux analysis. Despite its simplicity, the network is capable of generating flux distributions and ATP production rates that are comparable to a more detailed network. Second, HTMS makes use of microtiter well-plat technology and adaptations of well-sown enzymatic assays to increase precision and throughput for cell culture experiments. Multireplicate, multiparallel cultures in the sub-milliliter scale yield very precise metabolic rates using common laboratory equipment and at a fraction of the cost anal time of traditional experiments with T-flasks, spinner flasks, or bioreactor systems. The simplicity of he network and the well-plate assays synergistically comprise a new, extremely useful, broadly applicable, and relatively inexpensive way to probe cell cultures for metabolic effects, screen drugs and toxins, optimize media, and support the development of bioprocesses. The simplified network and cell culture and analytical assays are also useful for undergraduate, graduate, and professional training.
机译：为了在广泛的细胞培养应用中使用，我们设计了一种新颖的方法，称为高通量代谢筛选（HTMS），通过该方法可以更快地筛选哺乳动物细胞主要代谢途径的整体活性。该当前协议使用了来自代谢和细胞培养工程的理论和实验技术的改编。首先，HTMS雄性使用简化的代谢网络进行代谢通量分析。尽管其简单性，该网络仍能够产生与更详细的网络相当的通量分布和ATP生产率。其次，HTMS利用微量滴定板技术和对酶分析方法的完善来提高细胞培养实验的准确性和通量。使用通用的实验室设备，亚毫升级的多次重复，平行培养可产生非常精确的代谢速率，而使用T瓶，旋转瓶或生物反应器系统进行的常规实验的成本仅为肛门分析的一小部分。网络的简单性和酶标板的检测协同作用，构成了一种新的，极其有用的，广泛适用的且相对廉价的方法，可用于探测细胞培养物的代谢作用，筛选药物和毒素，优化培养基并支持生物过程的发展。简化的网络和细胞培养以及分析方法对于本科生，研究生和专业培训也很有用。

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