High-Speed Tandem Mass Spectrometric in Situ Imaging by Nanospray Desorption Electrospray Ionization Mass Spectrometry

Nanospray desorption electrospray ionization (nano-DESI) combined with tandem mass spectrometry (MS/MS), high-resolution mass analysis of the fragment ions (<i>m</i>/Δ<i>m</i> = 17 500 at <i>m</i>/<i>z</i> 200), and rapid spectral acquisition enabled simultaneous imaging and identification of a large number of metabolites and lipids from 92 selected <i>m</i>/<i>z</i> windows (±1 Da) with a spatial resolution of better than 150 μm. Mouse uterine sections of implantation sites on day 6 of pregnancy were analyzed in the ambient environment without any sample pretreatment. MS/MS imaging was performed by scanning the sample under the nano-DESI probe at 10 μm/s, while higher-energy collision-induced dissociation (HCD) spectra were acquired for a targeted inclusion list of 92 <i>m</i>/<i>z</i> values at a rate of ∼6.3 spectra/s. Molecular ions and their corresponding fragments, separated by high-resolution mass analysis, were assigned on the basis of accurate mass measurement. Using this approach, we were able to identify and image both abundant and low-abundance isobaric and isomeric species within each <i>m</i>/<i>z</i> window. MS/MS analysis enabled efficient separation and identification of isomeric and isobaric phospholipids that are difficult to separate in full-scan mode. Furthermore, we identified several metabolites associated with early pregnancy and obtained the first 2D images of these molecules