Ex Vivo Measurement of Stable Isotope-Labeled Fatty Acid Incorporation Into Phospholipids in Isolated Mice Muscle

With the advancement of liquid chromatography-mass spectrometry (LC-MS/MS), the quantification of glycerophospholipid (PL) molecules has become more accessible, leading to the discovery of numerous enzymes responsible for determining the acyl groups attached to these molecules. Metabolic tracer experiments using radioisotopes and stable isotopes are powerful tools for defining the function of metabolic enzymes and metabolic flux. We have established an ex vivo muscle experimental system using stable isotope-labeled fatty acids to evaluate fatty acid incorporation into PL molecules. Here, we describe a method to incorporate fatty acids with stable isotope labels into excised skeletal muscle and detect the PL molecules containing labeled acyl chains by LC-MS/MS. Key features • Quantify the metabolism of fatty acids into phospholipid acyl chains. • Enable measurements in excised muscle samples. • Assess the effects of genetic recombination of acyltransferases.