Measuring Extracellular Vesicles With a Nanoparticle Analyzer

Protocol Overview

The nanoparticles must be resuspended in liquid. In this case, extracellular vesicles (EVs) were resuspended in PBS at a 1:200 dilution factor. The re-suspension media must be confirmed as particle-free prior to use. 1ml of nanoparticles in suspension is then loaded into the pre-cleaned NanoSight visualization chamber. This is then fitted into place under the microscope and the laser module attached. The user must then manually locate a fixed point 'thumb print' under the microscope, and then align the field of view to the laser beam shining through nanoparticles, which can be found adjacent to the thumb print, just to the edge of the chrome line. The camera is then turned on and the settings adjusted to ensure the nanoparticles are well focused. Video captures are then recorded, in this case, 5 x 60s captures. This is achieved using the NanoSight software. The same software can then be used to process the captures and analyze the Brownian motion of the particles, thereby measuring their size distribution and concentration. All data, including a summary file, can be exported.