Applying flow cytometry-based isolation of platelet-derived microparticles to banked human plasma for cellular...

Platelet-derived microparticles (PMPs) constitute the majority of circulating microparticles in blood. PMPs carry cargo including RNA, protein, and miRNA, and play key pathophysiologic intercellular signaling roles in cardiovascular, autoimmune, and liver diseases. Most methods use fresh platelets treated with an agonist to generate PMPs, which has restricted the size and feasibility of human studies. With the rise of large biobanks, such as the UK Biobank (https://www.ukbiobank.ac.uk/), and because microparticles are retained in cryopreserved plasma, this study aimed to develop an efficient and reproducible method to isolate PMPs from banked cryopreserved human plasma to investigate their communication with target cells. We employed flow cytometry using 180-1,300 nm size calibration beads and the platelet-specific marker CD41 to identify and sort PMPs based on size and CD41 positivity. Following isolation, microparticle size and morphology were validated by electron microscopy and nanoparticle tracking analysis. To assess the utility of the microparticles for studies of cell-cell interactions, we visualized microparticle uptake into human umbilical vein endothelial cells and THP-1 cells. Our method enables the isolation and downstream analysis of human PMPs in cell-cell interactions, facilitating translational studies in large populations and rare diseases. © 2026 The Pathological Society of Great Britain and Ireland.