We describe the utility of intermediate-pressure MALDI and tandem mass spectrometry (MS/MS and MS n ) for the characterization and imaging of phospholipids in brain tissue sections. The use of both MS/MS spectra and MS/MS images allows for identification of isobaric compounds. The structural characterization of phosphatidylcholines, phosphatidylserines, phosphatidylethanolamines, and sphingomyelins directly from tissue sections is described.