colorimetric measurement of triglycerides cannot provide an accurate measure of stored fat content in drosophila比色测定甘油三酯不能提供果蝇储存的脂肪含量的准确测量.pdfVIP

Colorimetric Measurement of Triglycerides Cannot Provide an Accurate Measure of Stored Fat Content in Drosophila Bader Al-Anzi*, Kai Zinn* Division of Biology, California Institute of Technology, Pasadena, California, United States of America Abstract Drosophila melanogaster has recently emerged as a useful model system in which to study the genetic basis of regulation of fat storage. One of the most frequently used methods for evaluating the levels of stored fat (triglycerides) in flies is a coupled colorimetric assay available as a kit from several manufacturers. This is an aqueous-based enzymatic assay that is normally used for measurement of mammalian serum triglycerides, which are present in soluble lipoprotein complexes. In this short communication, we show that coupled colorimetric assay kits cannot accurately measure stored triglycerides in Drosophila. First, they fail to give accurate readings when tested on insoluble triglyceride mixtures with compositions like that of stored fat, or on fat extracted from flies with organic solvents. This is probably due to an inability of the lipase used in the kits to efficiently cleave off the glycerol head group from fat molecules in insoluble samples. Second, the measured final products of the kits are quinoneimines, which absorb visible light in the same wavelength range as Drosophila eye pigments. Thus, when extracts from crushed flies are assayed, much of the measured signal is actually due to eye pigments. Finally, the lipoprotein lipases used in colorimetric assays also cleave non-fat glycerides. The glycerol backbones liberated from all classes of glycerides are measured through the remaining reactions in the assay. As a consequence, when these assay kits are used to evaluate tissue extracts, the observed signal actually represents the amount of free glycerols together with all type