MS³ Imaging Enables the Simultaneous Analysis of Phospholipid CC and sn-Position Isomers in Tissues

Mass spectrometry (MS) imaging of lipids in tissues with high structure specificity is challenging in the effective fragmentation of position-selective structures and the sensitive detection of multiple lipid isomers. Herein, we develop an MS³ imaging method for the simultaneous analysis of phospholipid CC and sn-position isomers by on-tissue photochemical derivatization, nanospray desorption electrospray ionization (nano-DESI), and a dual-linear ion trap MS system. A novel laser-based sensing probe is developed for the real-time adjustment of the probe-to-surface distance for nano-DESI. This method is validated in mouse brain and kidney sections, showing its capability of sensitive resolving and imaging of the fatty acyl chain composition, the sn-position, and the CC location of phospholipids in an MS³ scan. MS³ imaging of phospholipids has shown the capability of differentiation of cancerous, fibrosis, and adjacent normal regions in liver cancer tissues.