Anti Sars-Cov Nonstructural Protein 13  

Immunofluorescence microscopy using Rockland's anti-SARS-CoV nsp13 antibody at 1:800, 6-h post infection Vero-E6 cells. For detection Cy3 conjugated Goat-anti-Rabbit IgG MX (611-104-122) was used. Personal Communication, Eric Snijder, Leiden University Medical Center, Leiden, Netherlands.

Colocalization of nsp13 with other replicase proteins and dsRNA in SARS-CoV-2-infected cells. (A) A549-ACE2 cells were infected with SARS-CoV-2 (MOI？=？2), fixed at 6-h postinfection (hpi), and costained with rabbit anti-SARS nsp13 pAb and appropriate home-made mouse anti-SARS-CoV-2 nsp sera or mouse anti-dsRNA MAb, followed by staining with Alexa Fluor 488-conjugated goat anti-rabbit Ab (green) and Alexa Fluor 555-conjugated goat anti-mouse Ab (red). Cell nuclei were stained with DAPI (blue) and examined by confocal microscopy. Images in the fifth column were obtained at higher magnification to show single-cell details of fluorescence labeling; scale bars represent 10？μm. The intensity distribution describes the colocalization of nsp13 with other replicase proteins for specific fluorescence along the indicated line. (B) Pearson’s correlation analysis demonstrated colocalization of nsp13 with other replicase proteins. One-way analysis of variance (ANOVA) was used for multiple comparisons on the colocalization between nsp13 and different nsps in GraphPad Prism 8.4.3 software. ****,？P？< 0.0001; ***,？P？< 0.001. Fig 8. PMID:？35730969

Time course of nsp8 and nsp13 detection. A549-ACE2 cells were mock infected or infected with SARS-CoV-2 (MOI 2), fixed at 2-, 3-, 4-, 6-, 8-, and 24-h postinfection (hpi), and costained with home-made mouse anti-SARS-CoV-2 N protein serum and rabbit anti-SARS nsp8 (A) or nsp13 (B) polyclonal, followed by staining with goat anti-mouse secondary antibody conjugated with Alexa Fluor 488 (green) and goat anti-rabbit secondary antibody conjugated with Alexa Fluor 555 (red). The cell nuclei were stained with DAPI (blue) and examined by confocal microscopy. No specific signal was observed at 2 and 3 hpi (not shown); scale bars represent 10？μm. The intensity distribution describes the timing of expression of nsp8 or nsp13 for specific fluorescence along the indicated line. (C) Pearson’s correlation was used to analyze changes in nsp8 or nsp13 over time. One-way analysis of variance (ANOVA) was used for multiple comparisons between different times among the nsp8 or nsp13 in GraphPad Prism 8.4.3 software. ****,？P？< 0.0001. Fig 6. PMID:？35730969