14b41 1 ap

Cultured cells were lysed with lysis buffer. Equal amounts of protein were run on 10 % SDS-PAGE and electrotransferred to polyvinylidene fluoride membranes (Millipore). After blocking with 5 % milk in TBST, membranes were incubated with primary antibodies overnight. The following antibodies were used: anti-ANT3 (1:1000, Proteintech, 14B41-1-AP), anti- FBLN1 (1:1000, Bioss, bs-0809R), E6/E7 (Abcam; ab70, ab30731) and anti-GAPDH (1:6000, Yeasen). Membranes were then incubated with the Rabbit peroxidase-conjugated secondary antibody (1:10,000, Abclonal). The blots were detected by sensitive chemiluminescence liquid (Yeasen), Biorad software was used to captured the images.

For immunofluorescence staining, Confocal dish (Nest, China) with planted cells were washed thrice with PBS and then incubated with 4 % Polyformaldehyde to fixation for 15 min. Then washed with PBS and treated with PBS containing 0.25 % Triton-X (Solabio, China) for20 min, blocked with 5 % Goat serum (Solabio, China) for 30 min in 37℃, and incubated with primary antibody (anti-ANT3, Proteintech, 14B41-1-AP), (anti- FBLN1, Bioss, bs-0809R) diluted in PBS overnight. Next, followed by washed thrice and incubated with proper secondary antibodies. Finally, the slides were incubated with DAPI, dehydrated, and mounted them for further evaluation in microscope.