FIG 4.

Dynapyrazole-A inhibits viroplasm fusion in the late stage of viroplasm formation. MA-104 cells were infected with RVA-NMTL (MOI = 0.1) and then treated with 3 μg/ml dynapyrazole-A and RVA-NMTL-neutralizing hyperimmune serum at 5 to 9 hpi. (A) Timeline of the experimental procedure. (B) Confocal microscopy was used to examine viroplasms (green) stained with a rabbit anti-NSP2 polyclonal antibody. Nuclei were stained with DAPI (blue). (C) The sizes of viroplasms were measured in 40 cells with Zen 2.3 software and analyzed statistically. (D) The numbers of viroplasms were analyzed statistically in three parallel experiments with 40 cells each time. Cells containing similar numbers of viroplasms were grouped, and the proportion of each group of cells of the total number of cells (40 cells) is the ordinate. (E) VP6, NSP2, NSP5, and GAPDH expression in the cell lysates was analyzed by WB with mouse anti-VP6 monoclonal antibody (1F4), rabbit anti-NSP2 polyclonal antibody, mouse anti-NSP5 monoclonal antibody (5E11), and rabbit anti-GAPDH polyclonal antibody, respectively. GAPDH was used as the loading control. Error bars represent the standard errors of the means (n = 3). *, P < 0.05; **, P < 0.01; ***, P < 0.001.