Recombinant SARS-CoV-2 pseudoviruses were produced by co-transfecting HEK-293T cells with 8.5 μg of pNL4-3ΔEnv-NanoLuc and 2.5 μg of pSARS-CoV-2-SΔ19 envelope spike plasmid as previously described . PEI (25 kDa linear, Polysciences, Inc., Warrington, PA) was the transfection reagent. At least 8 hours after transfection, the transfected HEK-293T cells were washed twice with DPBS (Thermo Fisher Scientific, Inc, Waltham, MA) and incubated with fresh media. After 48 hours of transfection, the supernatant was harvested and filtered using a 0.45 μm filter. The filtered supernatant was further purified using a 6-20% iodixanol gradient and centrifuged in an Optima LE-80k Ultracentrifuge with SW41 rotor (Beckman Coulter, Brea, CA) at 20,000 rpm for 2 hours at 4°C. Purified pseudoviruses (lower, heavier fractions) were collected and pooled together for infectivity testing and functional assays, including quantification of pseudovirus production by p24 content detection by ELISA.