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Measurement of long-chain fatty acid uptake into adipocytes.

Methods in enzymology (2014-02-18)
Elena Dubikovskaya, Rostislav Chudnovskiy, Grigory Karateev, Hyo Min Park, Andreas Stahl
ABSTRACT

The ability of white and brown adipose tissue to efficiently take up long-chain fatty acids is key to their physiological functions in energy storage and thermogenesis, respectively. Several approaches have been taken to determine uptake rates by cultured cells and primary adipocytes including radio- and fluorescently labeled fatty acids. In addition, the recent description of activatable bioluminescent fatty acids has opened the possibility for expanding these in vitro approaches to real-time monitoring of fatty acid uptake kinetics by adipose depots in vivo. Here, we will describe some of the most useful experimental paradigms to quantitatively determine long-chain fatty acid uptake by adipocytes in vitro and provide the reader with detailed instruction on how bioluminescent probes for in vivo imaging can be synthesized and used in living mice.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
N,N-Dimethylformamide, anhydrous, 99.8%
Sigma-Aldrich
Methanol, anhydrous, 99.8%
Sigma-Aldrich
Acetone-d6, "100%", 99.95 atom % D
Sigma-Aldrich
Ethylenediaminetetraacetic acid disodium salt dihydrate, suitable for electrophoresis, for molecular biology, 99.0-101.0% (titration)
Sigma-Aldrich
Sodium dodecyl sulfate, BioReagent, suitable for electrophoresis, for molecular biology, ≥98.5% (GC)
Millipore
NP-40 Alternative
Sigma-Aldrich
Dichloromethane, anhydrous, ≥99.8%, contains 40-150 ppm amylene as stabilizer
Sigma-Aldrich
Sodium deoxycholate, ≥97% (titration)