Anti fas

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).

Target cells were plated at 10,000 cells/well in Packard ViewPlate-96 and incubated 18–24 hours with and without the addition of IFNγ and TNFα (Roche) or resting or BiTE^®-activated T cells. Cells were fixed in 3.7% formaldehyde (ThermoFisher) and stained with anti-ICAM (Abcam) or anti-FAS (ThermoFisher) and Hoechst nuclear dye (ThermoFisher). Goat anti-mouse Alexa Fluor^® 488 or 647 secondary antibodies (ThermoFisher) were used for detection. Untreated cells were used to set fluorescence thresholds for percent positive.