Goat anti ace2

For SARS-CoV-2 spike subunit binding assay, cells were washed once with HBSS containing 2% FBS and incubated with 50 μg/mL S1-Fc, 200 μg/mL RBD-Fc (Sino biological, #40592-V08H) or NTD-Fc (Sino biological, #40591-V49H) for 30 min at 4°C, followed by washing two times with HBSS with 2% FBS. Cells were incubated with PE anti-human IgG for another 30 min at 4°C, washed two times, and fixed with 4% formaldehyde for 15 min. Cells were washed once, resuspended in HBSS with 2% FBS and analyzed by flow cytometry using FACSCelesta (BD Biosciences). To measure the surface expression of ACE2 and LRRC15, cells were washed with FACS buffer (1x PBS supplemented with 2% FBS and 1 mM EDTA) and stained with goat anti-ACE2 (R&D Systems, #AF933) at a 1:50 dilution or rabbit anti-LRRC15 (abcam, #ab150376) at a 1:100 dilution for 30 min at 4°C. Then the cells were washed two times and resuspended in FACS buffer containing the secondary antibodies at a 1:1000 dilution: AF647-labeled donkey anti-goat IgG (Invitrogen, #A32849) or AF488-labeled goat anti-rabbit IgG (Invitrogen, #A32731). After 30 min incubation at 4°C, the cells were washed two times, fixed with 4% formaldehyde for 15 min and washed and resuspended in FACS buffer before analyzing by flow cytometry using FACSCelesta (BD Biosciences) or Cytek Aurora spectral analyzer (Cytek Biosciences). Data was analyzed with Flowjo software.