Anti beta actin

Total proteins were extracted using RIPA buffer (Beijing Zoman Biotechnology Co., Ltd, China) supplemented with the protease inhibitor cocktail (Beijing Zoman Biotechnology Co., Ltd) according to the manufacturer's instructions. Proteins (20 μg) were separated by 10% SDS-PAGE (Biotides, China) and then transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, USA). The membranes were blocked in 5% milk for two hours and incubated with primary antibodies at 4°C overnight. Subsequently, the membranes were incubated with secondary antibodies at 37°C for one hour. Finally, proteins were visualized by using enzyme-linked chemiluminescence detection kit (ECL) and images were captured by a chemiluminescence imaging system (Azure Biosystems C300). The pixels from he western blot membranes were quantified by ImageJ software (NIH). Experiments were repeated three times.
NCM Universal Antibody Diluent was purchased from New Cell Molecular Biotech Co., Ltd. (China). Anti-mouse IgG antibody (H+L) (1:10000) and anti-rabbit IgG (H+L) antibody (1:10000) were purchased from Seracare (USA). Anti-beta actin (1:10000), anti-N-cadherin (1:1000), and anti-E-cadherin (1:1000) antibodies were purchased from GeneTex (USA). Anti-MMP-9 (1:1000) and anti-vimentin (1:5000) antibodies were purchased from Proteintech (China).

Cells were lysed in lysis buffer (50 mM Tris, 250 mM NaCl, 3 mM EDTA, 10% Triton X-100, 0.5% NP-40, and 10% glycerol). Cell lysate (10–50 μg) or viral supernatant (equal volume) were subjected to immunoblotting. Primary antibodies used in the study were anti-SARS-CoV-2 S2 (Genetex, Hsinchu, Taiwan; GTX632604), anti-VSV-M (Absolute, Boston, MA), and anti-beta-actin (Genetex, Hsinchu, Taiwan). The RBD antiserum was collected from rabbits immunized with RBD recombinant protein purified from E. coli. To remove N-linked oligosaccharides, viral supernatants were treated with peptide N-glycosidase F (PNGase F; NEB, Ipswich, MA) at 37°C for 1 h and then subjected to immunoblotting.