Authors:

Ricardo Pardo

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Verification of an Extracellular Vesicle Isolation Protocol from Blood Plasma Objective: Assessment of the Functionality of a Protocol for Isolating Extracellular Vesicles from Blood plasma Introduction Extracellular vesicles (EVs) are particles originating from cells, related to a wide range of physiological or pathological roles. Investigating EVs isolated from blood holds significant importance since blood participate in many physiological processes and EVs transport biological information. Methods: Firstly, arterial and venous punctures were performed to obtain blood samples. Subsequently, a two-step centrifugation process was employed to separate the EVs from other components of the blood. To further purify the EVs, size exclusion chromatography was conducted using qEV columns (qEVORIGINAL LEGACY, Izon Science Ltda. New Zealand) and 26 fractions were obtained. To determine the concentration of proteins, a BCA assay was performed, and nano sight was used to calculate the EV´s concentration. This comprehensive methodology ensures the efficient isolation and purification of EVs from blood, providing valuable samples for further analysis and investigation. Results: The equation y = 0.0003x + 0.6103 was obtained, resulting in a value of R was 0.9919. The protein concentration in arterial blood was 493.2203 μg/mL, while in venous blood it was 410.2203 μg/mL, indicating that the extracellular vesicles were collected in the expected fractions of their respective columns. In arterial blood the nano sight showed 1840012 particles/mL (30-130 nm), and 885494.8 particles/mL (>130 nm), showing the presence of EV`s. Conclusions: The essay confirmed the effectiveness of the employed protocol for isolating extracellular vesicles (EVs) from blood samples.

Source:

Purdue University / 2023

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Co-authors:

Ricardo Pardo