Journal: Virologica Sinica
Article Title: An inactivated recombinant rabies virus chimerically expressed RBD induces humoral and cellular immunity against SARS-CoV-2 and RABV
doi: 10.1016/j.virs.2022.12.006
Figure Lengend Snippet: Construction and identification of the recombinant viruses. A The construction scheme of the recombinant viruses SRV-nCoV-S1 and SRV-nCoV-RBD based on the reverse genetic operating system of RABV SRV9. B NA cells were infected with SRV-nCoV-S1, SRV-nCoV-RBD or wt-RABV SRV9. The cells were permeabilized or not at 48 h post-infection and then immunostained with antibodies against RABV G protein (red fluorescence) and SARS-CoV-2 RBD protein (green fluorescence). The nuclei were stained with DAPI (blue fluorescence). Scale bar: 10 μm. C The sucrose density gradient centrifugation-purified virions of recombinant viruses and wt-RABV SRV9 were analyzed by SDS-PAGE. The RABV proteins (RNA-dependent RNA polymerase (L), glycoprotein (G), nucleoprotein (N), phosphoprotein (P), and matrix protein (M)) and the S1 and RBD protein of SARS-CoV-2 were indicated on the corresponding bands. D Sucrose-purified virions were used to detect the expression of the exogenous proteins SARS-CoV-2 S1 and RBD. E Electron microscopy (EM) and dual-label immunogold EM detection of recombinant viruses. Gold particles (10 nm, red arrow) were used to label RABV G, and gold particles (18 nm, green arrow) were used to label the SARS-CoV-2 RBD protein. Scale bars represent 50 nm.
Article Snippet: The cells were blocked with 1% BSA for 30 min at 20∼25 °C) and incubated with a rabbit anti-SARS-CoV-2 RBD polyclonal antibody (PAb) (40592-T62, Sino Biological, 1:200) and mouse anti-RABV-G monoclonal antibody (mAb) (MAB8727, Abcam, 1:200) at 37 °C for 1 h. An FITC-conjugated anti-rabbit antibody (ab6717, Abcam, 1:300) and TRITC-conjugated anti-mouse antibody (T5393, Sigma, 1:500) were added and then incubated at 37 °C for 1 h. After staining with 4,6-diamidino-2-phenylindole (DAPI), the fluorescence was observed by confocal laser microscopy.
Techniques: Recombinant, Infection, Fluorescence, Staining, Gradient Centrifugation, Purification, SDS Page, Expressing, Electron Microscopy
HyTest is a verified supplier 
HyTest manufactures this product 
