Metabolic screening of mammalian cell cultures using well-plates.
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 makes 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-plate technology and adaptations of well-known 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 and time of traditional experiments with T-flasks, spinner flasks, or bioreactor systems. The simplicity of the 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.
Balcarcel
R Robert
RR
Department of Chemical Engineering, Vanderbilt University, VU Station B, Box 351604, Nashville, Tennessee 37235-1604, USA. r.balcarcel@vanderbilt.edu
Clark
Lindsey M
LM
D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation* / Q000379:methods*; D002478:Cells, Cultured; D020650:Combinatorial Chemistry Techniques / Q000295:instrumentation / Q000379:methods*; D003198:Computer Simulation; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods; D004734:Energy Metabolism / Q000502:physiology; D006801:Humans; D006825:Hybridomas / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D008322:Mammals; D008660:Metabolism / Q000502:physiology*; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000502:physiology; D011786:Quality Control; D020123:Sirolimus / Q000494:pharmacology
D009097:Multienzyme Complexes; D020123:Sirolimus