Tecnai g2 f30 field emission gun transmission electron microscope

To characterize the purified TEVs, we first used electron microscopy. TEVs suspended in PBS were placed on Formvar carbon-coated nickel grids. TEVs on grids were stained with 2% uranyl acetate and allowed to air dry. TEVs were visualized using an FEI Tecnai G2 F30 Field Emission Gun Transmission Electron Microscope with a 4k × 4k ultrascan charge-coupled device camera. The size distributions and concentration of TEVs isolated from cell culture supernatants were determined using the NanoSight LM-10 microscope (Malvern Instruments) equipped with particle tracking software. Ten independent microscopic fields were captured and analyzed per cell line sample. Data were merged and presented as a single histogram plot. In addition, we tested TEV-related protein markers from isolated TEVs using western blotting. TEV protein concentration was estimated by using the Pierce Micro BCA Protein Assay Kit. Ten micrograms of TEV protein from MC38 and modified cell lines were loaded on SDS gels. Primary antibodies binding markers: CD81 (1:1,000, BioLegend, cat. no. 104902) and ALIX (1:2,000, BioLegend, cat. no. 634502) were used to validate TEV protein markers. β-Actin (1:1,500, Cell Signaling, cat. no. 8H10D10) and β-tubulin (1:2,000, Invitrogen, cat. no. MA5 16308) confirmed no cellular contaminates.