Anti vimentin

Cell slides were blocked in goat serum for 15 min to block nonspecific antigens and then were incubated with anti-E Cadherin (ab76055, Abcam, Cambridge, MA, USA) and anti-PTPN2 (DF6629, AffinityBiosciences, Cincinnati, OH, USA) or anti-Vimentin (ARG57642, Arigo Biolaboratories, Wuhan, China) overnight at 4 °C. The next day, the slides were washed three times with PBS buffer, followed by either Alexa FluorR488 donkey anti rabbit IgG (H+L) or NovexR dropped on the slides, and incubated in an incubator at 37 °C for 30 min. Followed by counterstained with DAPI (5 mg/mL, Beyotime, Haimen, China), and then evaluated under afluorescence microscope (Nikon, Tokyo, Japan).

Cells were harvested and lysed with cell lysis reagent (NCM Biotech, Suzhou, China), and protein was collected. A total of 20 µg of total protein per lane was separated by 4-15% SDS-polyacrylamide gel electrophoresis and was then transferred to 0.2 μM or 0.45 μM PVDF membranes (Millipore, Billerica, USA). Subsequently, the membranes were blocked with 5% nonfat milk for 60-120 min. They were incubated separately at 4 °C overnight with the following primary antibodies: anti-BRD7 (Cat# 51009-2-AP, 1:500) and anti-GAPDH (Cat# 60004-1-Ig, 1:10,000) (Proteintech Group, Inc., Wuhan, China); anti-BIRC2 (Cat# 70008, 1:1,000), anti-E-cadherin (Cat# 14472, 1:1000) and anti-Snail (Cat# 3879, 1:1000) (purchased from CST, MA, USA); anti-Flag (Cat# F2555, 1:1000, Sigma-Aldrich, St. Louis, MO); anti-Vimentin (Cat# ARG66199, 1:1000, Arigo, Taiwan, China) and anti-N-cadherin (Cat# AF5239, 1:1000, Affinity Biosciences, Jiangsu, China). The membranes were then incubated with species-matched secondary antibodies for 60 min at 37 °C. Bands were visualized by Western Blotting Substrate (Share-Bio, Shanghai, China), and signals were detected by an enhanced chemiluminescence detection system according to the manufacturer’s protocol (MiniChemi™ I, SAGECREATION, China).