Flow Cytometric Analysis of SARS-CoV-2 Infection

Cells were dislodged by washing in PBS and treatment with 0.05% trypsin (Gibco) for 20 min, followed by pipetting and centrifugation. For all subsequent steps, reagents were diluted in FACS buffer (PBS, 1% FCS, 2 mM EDTA) unless stated otherwise, and washed twice with FACS buffer between steps. Cells were stained with Live/Dead Fixable Aqua Cell Stain (1:200 in PBS, Life Technologies, L34957) combined with FcR block (1:200 in PBS, eBioscience), fixed in 4% formaldehyde (10 min at room temperature), and permeabilised in PBS containing 0.1% Triton-X (20 min at room temperature). Cells were incubated with antibodies against human MxA (clone M143, kind gift from G Kochs) and SARS-CoV-2 N protein (clone EY-2A, kind gift from Alain Townsend^{59 (link)}; 1:200, 30 min, 4 °C), and goat anti-mouse AlexaFluor 488 (Life Technologies, A11029) and anti-human AlexaFluor 647 (1:500, 30 min, 4 °C; Life Technologies, A21445), and resuspended in CellFix (1:10 in water; BD, 340181). Cells were analysed by flow cytometry on an Attune NxT Flow Cytometer (Thermo Fisher Scientific) and data were analysed using FlowJo software (BD).