Skip to main content
. 2021 Mar 16;184(9):2316–2331.e15. doi: 10.1016/j.cell.2021.03.029

Figure 3.

Figure 3

Virus neutralization and binding of infected cells by anti-SARS-CoV-2 mAbs targeting the NTD

(A) SARS-CoV-2-infected Vero cells were stained with serial dilutions of COV2-2489, COV2-2676, COV2-2381, or DENV2-2D22 (isotype control) prior to analysis of staining intensity by flow cytometry. The positively stained cells were gated using uninfected and isotype control mAb-stained infected cells, and the integrated mean fluorescence intensity (iMFI) was determined by MFI of positive cells multiplied by the percent of total positive cells. The left panel shows representative dose response curves for the staining intensity of infected cells by each mAb. The right panel shows the mean EC50 values for infected cell staining, determined from three independent experiments. Error bars represent SEM.

(B–D) The neutralization potency of COV2-2489 and COV2-2676 against SARS-CoV-2 was assessed by FRNT using (B) Vero, MA104, 293T+ACE2, and Vero+TMPRSS2 cells. Results are representative of three independent experiments performed in duplicate. (C) COV2-2489 and COV2-2676 were assayed for neutralization potency by modified FRNT in which mAb was added to SARS-CoV-2 before (pre-attachment, filled circles) or after (post-attachment, open circles) virus was absorbed to Vero E6 cells. (C) Error bars represent the range from two technical replicates. Data shown are representative of three independent experiments. (D) Attachment inhibition. (Left panel) Vero or Vero+hACE2+TMPRSS2 cells were incubated with SARS-CoV-2 at 4°C for 1 h. After extensive washing, cell-bound viral RNA was measured by qRT-PCR. (Right four panels) SARS-CoV-2 was pre-incubated with 5 or 50 μg/mL of indicated anti-RBD or anti-NTD mAbs for 1 h prior to addition to Vero or Vero+hACE2+TMPRSS2 cells. Cell-bound viral RNA was measured by qRT-PCR. Data are pooled from three independent experiments. (Left) t test: ∗∗∗∗p < 0.0001; (Right) one-way ANOVA with Dunnett’s multiple comparisons test compared to isotype control mAb treatment: ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

(E) Neutralization curves for COV2-2196 IgG, F(ab′)2, F(ab), or rDENV-2D22; COV2-2489 IgG, F(ab′)2, or F(ab); or COV2-2676 IgG, F(ab′)2, or F(ab) in a SARS-CoV2-rVSV assay using RTCA. Error bars indicate SD; data represent at least two independent experiments performed in technical triplicates.