High-Throughput In-Depth Single-Cell Lipidomics by Ozone-Induced Dissociation and Mass Cytometry

Lipids are essential chemical components of living cells, and performing in-depth lipidomics at the single-cell level provides profound insights into the cellular structure and function. Here, we couple ozone-induced dissociation (OzID) with mass cytometry to develop a high-throughput in-depth single-cell lipidomics platform. By precisely controlling the OzID reaction conditions to cleave approximately half of the C═C bonds in unsaturated lipids, appropriate abundances of diagnostic products are generated while retaining a comparable proportion of unreacted lipids. Subsequently, high-throughput single-cell mass cytometry analysis is performed to obtain in-depth lipidomic information comprising intact lipids and C═C structural information. The in-depth lipidomic platform achieves a throughput of 166 cells/min, enabling clear discrimination among clinically relevant cell types with highly similar intact lipid profiles, e.g., normal lung epithelial cells (Beas-2B) versus non-small-cell lung cancer cells (A549 and H1299). Our platform demonstrates outstanding potential for discriminating cell types and discovering refined lipid biomarkers.