2.3 Confocal microscopy assay
HeLa cells (2×105) were seeded on glass slides 10 h before transfection. The HeLa cells were transfected with Lipofectamine 3000 reagent (Thermo Fisher). Twenty-four hours later, the cells were washed, fixed with 4% paraformaldehyde, and permeabilized with 0.1% Triton X-100. After washing, the cells were blocked with 5% bovine serum albumin (BSA). Cells were incubated with an anti-HA primary antibody for 1 h at RT for the immunofluorescence assay. After 3 washes, cells were stained with an Alexa Fluor 647-conjugated secondary antibody for 1 h at RT. Following 3 washes with PBS, cells were stained with DAPI for 30 min, washed for 1 h at RT, and used for confocal microscopy assay (ZEISS LSM700). At least 100 random cells per slide were analyzed, and the most representative images from each slide were selected for presentation.