Single-Cell Structural Lipidomics Using Miniature Dual-LIT Mass Spectrometer

Abstract

Structural lipidomics provides comprehensive information of lipidomes and it is of a significant interest in applying it to single-cell analysis for accurate cell phenotyping and lipid pathway studies. However, structural lipidomics relies on tandem mass spectrometry analysis of a large number of lipid species, which remains challenging for single-cell analysis due to the small sample amounts. Here, using a miniature dual-LIT (linear ion trap) mass spectrometer with enhanced capability of ion processing, we developed an effective strategy to achieve annotations of more than 100 lipid species in a single cell with high structural specificity. Ion utilization efficiency was greatly improved with multi-stage MSn (n = 2-4) analysis performed for each lipid species to acquire structure information at different levels. From a single MDA-MB-468 cell, we identified 100+ lipids, including 64 lipids at the acyl-chain sum composition level, 23 at the sn-position level, 30 at the C=C location level, and 20 at the C=C/sn-position level, primarily phosphatidylcholamines (PCs) and phosphatidylethanolamines (PEs). Significant variations in sn-position and C=C location isomers were observed in Doxorubicin-resistant and -sensitive K562 cells. With enriched information at the structural lipidomics level, the correlation was established between variations in lipidome and response to ferroptosis for human cancer cells.