J. Lipid Res.
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A more recent version of this article appeared on April 1, 2007

Papers In Press, published online ahead of print January 9, 2007
J. Lipid Res., doi:10.1194/jlr.D600047-JLR200
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Submitted on December 4, 2006
Revised on January 8, 2007
Accepted on January 8, 2007

Cell based multiwell assays for detection of substrate accumulation and oxidation

Andreas J. Wensaas, Arild C. Rustan, Karina Lövstedt, Bengt Kull, Sven Wikström, Christian A. Drevon, and Stefan Hallén

Molecular Pharmacology, AstraZeneca R&D CVGI Mölndal, Mölndal 43183

Corresponding Author: stefan.hallen{at}astrazeneca.com

We describe multiwell assays for detecting accumulation as well as subsequent oxidation of 14C-labeled substrates in cultured cells. Accumulation is monitored in real time by an established scintillation proximity assay (SPA) where the scintillator is embedded in the plate base primarily detecting cell-associated radiolabel. The substrate oxidation assay is a novel variant of previously described experimental approaches aimed at trapping 14CO2 produced by isolated enzymes, organelles or intact cells. This method uses a standard 96 well tissue culture plate and on top, an inverted filter plate immersed with NaOH that are clamped into a sandwich sealed with a silicon gasket to obtain gas-tight compartments. 14CO2 is captured in the filter and quantified by conventional scintillation. We demonstrate both accumulation and subsequent oxidation of 14C-labelled substrates in cultured human myotubes, adipocytes and hepatocytes. Both methods are adaptable for compound screening and at the same time these protocols provide easy-to-use and time- saving methods for in vitro studies of cellular fuel handling.


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D. M. Schwartz and N. E. Wolins
A simple and rapid method to assay triacylglycerol in cells and tissues
J. Lipid Res., November 1, 2007; 48(11): 2514 - 2520.
[Abstract] [Full Text] [PDF]


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