Fig. 3.

Probing T3 reovirus binding to sialylated glycans on living cells. a Combined optical microscopy and FD-based AFM of T3SA+ binding to cells expressing (CHO) or lacking (Lec2) α-SA on the cell surface. b Overlay of DIC, eGFP, and mCherry signals of a confluent layer of co-cultured fluorescent CHO cells (actin-mCherry and H2B-eGFP) and Lec2 cells. c, d FD-based AFM topography image (c) and corresponding adhesion map (d) from probing of adjacent cells indicated in the dashed square in b. The adhesion map shows interactions mainly on CHO cells (α-SA-expressing cells) (white pixels). For higher visibility, the pixel size in the adhesion image was enlarged two-fold. e DFS plot of data from α-SA model surfaces (grey circles, from Fig. 2c) and living cells (red dots). Histogram of the force distribution observed on cells fitted with a multi-peak Gaussian distribution (n = 700 data points) is shown at the side. f Box plot of BF observed for T3SA+ (gray) and T3SA− (white) virions as well as T3SA+ virions following injection of 1 mM Neu5Ac (red). The horizontal line within the box indicates the median, boundaries of the box indicate the 25th and 75th percentile, and the whiskers indicate the highest and lowest values of the results. The square in the box indicates the mean. For all experiments, data are representative of at least n = 5 independent experiments. g Influence of SA on virus binding determined by flow cytometry. Cells were incubated with either PBS (Mock) or Alexa Flour 488-labeled T3SA+ or T3SA− virions (10⁵ particles per cell), and the median fluorescence intensity (MFI) of cell-bound virus was determined by flow cytometry as shown in Supplementary Fig. 3a. Error bars indicate s.d. of the mean value. Experiments were repeated twice (n = 2 independent experiments, each with duplicate samples). ns, P > 0.05; ****P < 0.0001; determined by two-sample t-test in Origin (f) and by two-way ANOVA corrected for multiple comparisons using Tukey’s test in GraphPad Prism (g), respectively. Source data are provided as a Source Data file