Using a uniform design strategy [37,38], we optimized the transfection ratio of the S, M/E, N and Fluc-PS9 plasmids, resulting in an approximately fourfold increase in infection signal compared to previously reported conditions [36] (S1B Fig).Omission of any structural proteins completely abolished the SC2-VLP infectivity, which remained strictly dependent on ACE2 and TMPRSS2 expression (S1C Fig and S2 Table). Here, ACE2 is linked to infection.