Optimised Sample Preparation for Metabolic Analysis of Cell Cultures Using DESI-MS

Abstract

Desorption electrospray ionisation mass spectrometry (DESI-MS) can profile cultured cells with minimal sample preparation, but salt carryover and wash-induced leakage often reduce metabolite coverage and reproducibility. Using the non-adherent human Burkitt’s lymphoma B cell line Ramos, we evaluated different substrates. Reversed-phase C18 thin-layer chromatography (TLC) plates produced homogeneous and robust signals of the deposited cellular material, while enabling passive separation of soluble contaminants from the cell-dense region. We then optimised wash solvents and frequency. Ammonium acetate (AA) washes are frequently used by the community to replace phosphate-buffered saline (PBS) washes. However, we observed that repeated AA washes markedly reduced cell viability, whereas PBS washing left residual salts that suppressed ion signals. A single 1:1 PBS/AA wash best balances desalting with cell integrity and improved coverage and reproducibility across small polar metabolites and lipids. Finally, we applied the workflow to human pancreatic ductal adenocarcinoma cell line, PANC-1, treated with a glutaminase 1 inhibitor and captured glutamine accumulation with depletion of glutamate and downstream metabolites. This rapid, extraction-free molecular profiling approach supports high-throughput phenotyping and can help reveal metabolic diversity and candidate biomarkers across cell models. Repeated measurements on the same sample support screening-scale metabolic phenotyping and the recording of additional acquisition modes.