2.5. Western Blot Analysis
HT
DCTM protein assay (Bio-Rad, Hercules, CA, USA) was used to measure the protein content of exosome samples in the microplate reader Synergy HT (BioTek, Winooski, Vermont, USA). Samples containing 40 μg of protein were fractionated by SDS-PAGE on 12% polyacrylamide gels. Then, separated proteins were transferred to PVDF membranes and pre-incubated in 5% non-fat milk for 30 min at 37 °C to block non-specific binding. After that, membranes were incubated overnight at 4 °C with specific primary antibodies. Antibodies against Flotillin-1 (Ref. sc-20066), CD63 (Ref. sc-5275), CD81 (Ref. sc-166029), CD9 (Ref. sc-13118), CD14 (Ref. sc-9150), VDAC1 (Ref. sc-8828) and LAMP-1 (Ref. sc-5570) were purchased from Santa Cruz Biotechnology, Dallas, TX, USA; LAMP2A (Ref. ab18528), HSP70 (Ref. ab2787) and HSP60 (Ref. ab1428) were purchased from Abcam