This paper describes the application of nanoparticle bombardment with time‐of‐flight secondary ion mass spectrometry (NP‐ToF‐SIMS) for the analysis of native biological surfaces for the case of sagittal sections of mammalian brain tissue. The use of high energy, single nanoparticle impacts (e.g. 520 keV Au 400 ) permits desorption of intact lipid molecular ions, with enhanced molecular ion yield and reduced fragmentation. When coupled with complementary molecular ion fragmentation and exact mass measurement analysis, high energy nanoparticle probes (e.g. 520 keV Au 400 NP) provide a powerful tool for the analysis of the lipid components from native brain sections without the need for surface preparation and with ultimate spatial resolution limited to the desorption volume per impact (~10 3 nm 3 ). Copyright © 2012 John Wiley & Sons, Ltd.