β actin antibody clone ac 74

VeroE6 cells were seeded in 6-well plates and infected with BA.2 or BA.1 at 0.1 MOI. Cell lysates were harvested in 200 μL RIPA buffer (89901, Thermo Scientific ) at 24 h post infection for the analysis of spike processing. The samples were subjected to 8% of SDS-PAGE and transferred to the PDVF membranes, followed by blocked with 5% skim milk in PBS for 2h at room temperature and incubated with specific primary antibodies at 4°C overnight, followed by incubating with horseradish peroxidase (HRP) conjugated secondary antibodies (Thermo Fisher Scientific) for 1h at room temperature. The signal was developed using SuperSignal West Pico PLUS Chemiluminescent Substrate (34580, Thermo Scientific, USA) and detected using Alliance Imager apparatus (Uvitec, Cambridge, UK). Full-length spike and S2 was detected with a rabbit anti-SARS-CoV-2 spike S2 antibody (40590-T62, Sino Biological) (1:5000). Nucleocapid (N) was detected with an in-house rabbit anti-SARS-CoV-2 N immune serum (1:10000) and β-actin was detected with a β-actin antibody (clone AC-74, A5316, Sigma, USA) (1:5000).

VeroE6-TMPRSS2 cells were challenged with Omicron subvariants at 0.5 MOI and lysed in RIPA buffer (89901, Thermo Fisher Scientific) at 48 hpi for Western blot analysis. The membranes were blocked with 5% milk for 2 h at room temperature and incubated with a rabbit anti-SARS-CoV-2 spike S2 antibody (40590-T62, Sino Biological, China) at 4 °C for overnight incubation, followed by detection with horseradish peroxidase (HRP) conjugated secondary antibodies (31460, Thermo Fisher Scientific) for 1 h at room temperature. Antigen signal was developed using SuperSignal West Pico PLUS Chemiluminescent Substrate (34580, Thermo Scientific) and detected using an Alliance Imager apparatus (Uvitec, Cambridge, UK). β-actin was detected with a β-actin antibody (clone AC-74, A5316, Sigma-Aldrich) (1:5000).