Together with the International Society on Thrombosis and Haemostasis (ISTH), the European Metrology Research Programme METVES, and the Vesicle Observation Center of the Amsterdam University Medical Centers, Exometry initiated a project to standardize extracellular vesicle (EV) measurements by flow cytometry. The project ran from 2014 to 2018.
Reliable and standardized detection is a prerequisite for the utilization of EVsextracellular vesicles as biomarkers for disease. Flow cytometers have the potential to reliably count and identify single EVsextracellular vesicles at a rate of thousands per second. However, because flow cytometers differ in optical configuration, different flow cytometers obtain different EVextracellular vesicle concentrations for the same sample. Moreover, flow cytometers provide data in arbitrary units, thereby hampering data comparison between flow cytometers and laboratories.
The goal of this standardization study was to improve the reproducibility of the measured concentration of EVsextracellular vesicles by different commercially available flow cytometers.
To standardize EVextracellular vesicle measurements, flow cytometers were calibrated using traceable reference materials and light scattering theory. The used reference materials have been extensively characterized by European metrology institutes in the METVES project. Exometry has developed easy-to-use software to automatically recognize the reference materials, obtain the optical configuration of the flow cytometer, and set an EVextracellular vesicle size gate in nanometers. This size gate is calculated with Mie theory and takes into account the optical configuration of the flow cytometer and the refractive index of EV. The size gate ensures that each flow cytometer will determine the concentration of EVextracellular vesicle in the same size range.
We kindly thank the Scientific Standardization Committee on Vascular Biology of the ISTH for funding this project.
The project has resulted in a scientific publication: