Nano-CET (Dispertech) was used to determine the refractive index (RI) of extracellular vesicles (EVs)<100 nm, as it simultaneously measures the trajectory and scattering intensity of particles within a nanofluidic optical fiber. A test sample of outdated erythrocyte-derived EVs was developed for RI determination. Polystyrene beads and silica beads of known sizes were used for calibration and validation. A Python algorithm was developed to track the position and scattering intensity of single particles. The determined RIs of silica beads and hollow organosilica beads (HOBs) matched literature values. Measured RI of EVs was higher (1.63±0.06) than anticipated (RI~1.48), perhaps due to an elevated hemoglobin concentration in outdated erythrocyte EVs.
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