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Direct coupling of high-performance thin-layer chromatography (HPTLC) to matrix-assisted laser desorption/ionization quadrupole ion trap time-of-flight mass spectrometry (MS) was shown to be a reliable and reproducible method to obtain structural information and fundamental properties of glycosphingolipids (GSLs).We report a protocol for the preparation of neutral GSL extracts frommouse tissues and demonstrate the applicability of HPTLC/MS to these preparations. The protocol consists of lipid extraction and ion exchange chromatography followed by a mild alkaline treatment and a reversed-phase cartridge extraction. Possible structures for each GSL are proposed based on HPTLC/MS analyses. This fast and simple method can be used to screen neutral GSL extracts obtained from tissues and cells without isolation and purification into individual GSLs.

High-Performance Thin-Layer Chromatography/Mass Spectrometry for Rapid Analysis of Neutral Glycosphingolipids