Exosome Block Preparation: A Protocol to Preserve Exosomes From Cultured Human Colon Cancer Cells

Abstract

Source: Jung, M. K. et al. Sample Preparation and Imaging of Exosomes by Transmission Electron Microscopy. J. Vis. Exp. (2018)

This video describes the technique of preparing exosome blocks from cultured gastrointestinal tumor cells. These exosome blocks can further be sliced, stained, and imaged to determine the position of specific proteins in the lipid bilayer vesicle.

Protocol

1. Block Preparation of Exosomes Pellet the exosomes from the culture supernatant of HCT116 cells by centrifugation at 100,000 x g for 1.5 h. Remove the culture supernatant and carefully fix the purified exosome pellet with 1 mL of 2.5% glutaraldehyde in 0.1 M sodium cacodylate solution (pH 7.0) for 1 h at 4 °C. For 0.1 M sodium cacodylate, dissolve 4.28 g cacodylic acid in 160 mL of distilled water (DW). Adjust pH to 7.4 with 0.1 M HCl, then make up to 200 mL with distilled wat…

Materials

Glutaraldehyde	EMS	16200
Sodium cacodylate	EMS	12300
Osmium tetroxide 1 g crystal	EMS	19100	Use only in fume hood
Acetone	JUNSEI	1B2031