Flow Cytometry Has a Significant Impact on the Cellular Metabolome

The
characterization of specialized cell subpopulations in a heterogeneous
tissue is essential for understanding organ function in health and
disease. A popular method of cell isolation is fluorescence-activated
cell sorting (FACS) based on probes that bind surface or intracellular
markers. In this study, we analyze the impact of FACS on the cell
metabolome of mouse peritoneal macrophages. Compared with directly
pelleted macrophages, FACS-treated cells had an altered content of
metabolites related to the plasma membrane, activating a mechanosensory
signaling cascade causing inflammation-like stress. The procedure
also triggered alterations related to energy consumption and cell
damage. The observed changes mostly derive from the physical impact
on cells during their passage through the instrument. These findings
provide evidence of FACS-induced biochemical changes, which should
be taken into account in the design of robust metabolic assays of
cells separated by flow cytometry.