Tecnai g2 20 s twin tem

MDA-MB-231 EVs (10⁸ μL⁻¹) before VMF were adsorbed on Formvar/carbon-coated copper grids for 20 min. MDA-MB-231 EVs after VMF ( ~ 8 × 10⁶ μL⁻¹) were adsorbed on Formvar/carbon-coated copper grids for 1.5 h. The adsorbed EVs were then stained by 5 drops of 3% uranyl acetate. After drying, the samples were observed on a Tecnai G2 20 S-TWIN TEM (FEI, USA) at 200 kV.

25 ml parasite culture was grown to a density of ~20 × 10⁶ cells/ml to obtain a reasonably large pellet for TEM. The culture medium was washed twice with HBSSDG at 800 × g for 5 min. For live (non-demembranated) cells, the final washed pellet was fixed in 2.5% glutaraldehyde and 2% paraformaldehyde in phosphate buffered saline for 2 hours at room temperature followed by 4 °C overnight. For demembranated cells, the washed pellet was demembranated in LRP medium containing 0.1% Triton X-100 (without ATP) for ~30 seconds, after which the demembranated cells were pelleted down and fixed as mentioned above. Before fixation, both live and demembranated cells were checked under the microscope to ensure proper removal of media that interfere with TEM imaging, and to ensure adequate demembranation of the treated cells. Blocks were prepared, sectioned and stained at SAIF, AIIMS, New Delhi. Electron micrographs were captured on a Tecnai G2 20 S-Twin TEM (FEI) and captured with Gatan, 4 k × 4 k camera at SAIF, AIIMS.