Anti sars cov 2 np

Sections were incubated at 4°C overnight with primary mouse originated antibodies including anti-SARS-CoV-2 NP (#ab273434, 1:500, mouse IgG1; Abcam), anti-ACE2 (#MA5-31395, clone ID: CL4035, 1:400, mouse IgG1; Invitrogen), anti-CD68 (#MAB11303, clone KP1, 1:300, mouse IgG1; Bio-RAD), anti-CD169 (#346020, clone ID: 7-239, 1:200, mouse IgG1; Biolegend), anti-B220 (#MAB11301, clone ID: 123C3, 1:100, mouse IgG1; Bio-RAD), anti-CD11c (#ab254183, clone ID: KB90, 1:200, mouse IgG1; Abcam), and anti-Fas (#48095942, 1:200, mouse IgG1; ThermoFisher), and rabbit originated antibodies including anti-SARS-CoV-2 NP (#clone ID: 019, 1:200, rabbit IgG; Sino Biological), anti-IL-1β (#ab9722, rabbit IgG1, Abcam) and anti-IL-6 (#12153, Rabbit mAb, CST). After washing with PBS (3 washes, 5 min per wash), sections were incubated with Alexa Fluor^® 555-conjugated goat anti-rabbit IgG antibodies (#ab150078, 1:200, Abcam) or Alexa Fluor^® 488-conjugated goat anti-mouse IgG1 antibodies (#ab150078, 1:200, Abcam) for 1 h. Finally, the sections were incubated with 1 μg/ml DAPI (Sigma, St. Louis, MO, USA) for 10 min to stain the nuclei. Sections incubated with the appropriate isotype control antibodies and fluorescently labelled secondary antibodies were used as negative controls. Results were analyzed using fluorescence microscopy (Zeiss Axioplan 2).

Western blot analysis was performed according to standard procedures. Protein samples were separated by 10% SDS-PAGE, blotted onto Protran-BA83 nitrocellulose membranes (Schleicher & Schuell, Keene, NH, USA) and incubated overnight with mouse monoclonal anti-SARS-CoV-2 Np (1:1000) (SinoBiological, Eschborn, Germany), polyclonal rabbit anti-DDX3X A300-474A (1:1000) (Bethyl, Montgomery, TX, USA) and polyclonal rabbit anti-DDX1 (1:1000) (Bethyl, Montgomery, TX, USA), followed by 1 h of incubation with HRP-conjugated secondary antibodies known as goat anti-mouse or goat anti-rabbit (1:5000) (Sigma, St. Louis, MO, USA). Chemiluminescent signals were developed using the ECL reagent (GE Healthcare, Uppsala, Sweden) and Chemidoc imaging system (Bio-Rad, Hercules, CA, USA).