 Membrane vesicles are key vehicles for intracellular and intercellular transport. Extracellular vesicles are released from cells by several mechanisms including secretion from multivesicular bodies (exosomes) and shedding from the cell surface (ectosomes, aka microvesicles or microparticles). EVs can bear cellular surface markers and cargo but, being ~100 times smaller than a cell, have ~10,000 times less surface area, and ~1,000,000 times less volume, so the abundance of vesicle surface markers and intra-vesicular cargo will be much lower than the cell of origin. |
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Flourescence-based Detection and Sizing of EVs While light scatter-based detection of cells in routine using flow cytometry, EV’s small size make light scatter-based detection difficult. Moreover, it can be challenging to accurately estimate EV size from light scatter intensity. We have developed EV staining, detection, and calibration protocols that enables the efficient detection and sizing of individual EVs as small as ~70 nm in diameter, directly in complex biofluids such as plasma and serum, without the need for isolation or purification. |
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Sensitive Detection of EV Surface Markers and Cargo Because EVs may have surface areas that are several orders of magnitude less than the cell of origin, even highly abundant cell surface molecules may be present at less than 100 copies per EV, challenging the detection limit of conventional flow cytometers. Moderate and low abundance surface markers may be below the limit of detection. Using high sensitivity flow cytometers, optimized staining and measurement protocols, and appropriate instrument and assay calibration procedures, we are making sensitive and quantitative measurements of EV surface markers. |
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Selected References: Flow Cytometry of Extracellular Vesicles: Potential, Pitfalls, and Prospects A Trigger Channel Threshold Artifact in Nanoparticle Analysis High Sensitivity Flow Cytometry of Membrane Vesicles |
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