Fig. 4

Lactadherin protein is present in BC cell-secreted sEVs. sEVs secreted by different human BC cell lines were obtained through cell culture conditioned medium ultracentrifugation and the number, size and integrity were analyzed by A Nanotracking analysis and B Transmission electron microscopy (representative images of sEV-MCF10A (up) and sEV-MDA231 (down)). C Detection of sEV markers in human BC cell-secreted sEVs. Total protein extracts were loaded in 10–12% acrylamide gels and transferred to nitrocellulose membranes. CD63, TSG101 and Flotillin-1 were used as sEV markers. Calnexin was used as intracellular contamination protein. Human normal mammary epithelial MCF10A cells was used as control. D Lactadherin protein level was measured in sEVs through an ELISA assay. Fifty microliters of each sEV per well were used in the assay. Results were normalized against 100 µg total protein amount, quantified by microBCA total protein quantification assay. The graph shows the mean ± SD. Images and data are representative of three independent experiments. * p < 0.05; ** p < 0.01