Plasma membrane order maps functional diversity in immune cells.

Cell membranes undergo biophysical remodeling as an adaptation to the surroundings and to perform specific biological functions. However, the extent and relevance of such changes in human immune cells remain unknown, largely because of the lack of single-cell and multidimensional methodologies. Here we apply a cytometry-based method to fill this gap by combining biophysical profiling with simultaneous analysis of immune cell markers. This platform reveals notable cell-type-dependent plasma membrane order heterogeneity in immune cells. By sorting immune cells according to their membrane order and performing transcriptome and spatial surface proteome analyses together with functional tests, we show that plasma membrane order can be used to identify subsets of immune cells with distinct phenotypes and functional behaviors. Our findings demonstrate a broad heterogeneity of plasma membrane order in immune cells that will provide a more precise definition of immune cell states on the basis of their biophysical properties in health and disease.