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. 2023 Jun 23;299(8):104955. doi: 10.1016/j.jbc.2023.104955

Figure 7.

Figure 7

ORF8 limits the reactivity of anti-SARS-CoV-2 (convalescent or vaccinated) human sera towards Spike-producing cells.A–C, HEK293T cells cotransfected with a plasmid encoding Spike-Flag and a bicistronic plasmid encoding mCherry and ORF8-Strep genotypes (ORF8-Strep, ORF8-Strep S84L) were harvested and incubated with anti-SARS-CoV-2 human sera collected from three COVID-19 convalescent (A and B), six vaccinated (C) (three Pfizer and three Moderna, before first shot (prevaccination) and after second shot (postvaccination)), or three COVID-19 negative donors (A). The IgG molecules in the sera that reacted to Spike-producing cells were fluorescently labeled using antibodies that specifically recognize the Fc region of human IgG molecules. The human IgG Fc levels in viable (LIVE/DEAD-negative) cells expressing ORF8-Strep (mCherry-positive) were evaluated by flow cytometry. The data are combined from three independent experiments and represented as mean ± s.d. Statistical significance was analyzed using one-way (B, Dunnett’s test) or two-way ANOVA (C, Tukey’s test). Fc, fragment crystallization; IgG, immunoglobulin G; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.