Coligation of LFA-1 is required for influenza viral particles to trigger NK cell symmetric spreading. (A) Representative bright-field, IRM, and fluorescence images of pNK cells, stained with phalloidin–AlexaFluor-488, stimulated on surfaces coated with a low (400 HU/mL; left panels) or high (4000 HU/mL; right panels) concentrations of influenza virus particles, with or without ICAM-1. Bars represent 10 μm. (B) Graph represents the proportion of pNK cells that form a peripheral ring of F-actin when influenza virus particles were titrated onto glass surfaces coated with poly-L-lysine with or without ICAM-1. Graph represents mean ± SEM, from 4 independent experiments counting > 70 cells per condition. (C) Quantification of the pNK cell surface contact areas determined by IRM on slides coated with ICAM-1 and increasing concentrations of influenza virus particles (0-4000 HU/mL). Data are mean ± SEM for n > 300 cells per condition from 4 independent experiments. (D) The symmetry of the contact interface for cells stimulated on slides coated with ICAM-1 and increasing concentrations of influenza virus particles is represented as the SD of the radial distances. Data represent the distance from the NK cell center to the circumference measured at 360 radii for n = 87-99 cells, from 3 independent experiments. Graphs represent the median ± SE of the median for each dataset. ***P < .0001. *P < .01. NS indicates not significant, analyzed by 1-way ANOVA. (E) The SD of radial distances was compared for pNK cells, from 3 independent donors, stimulated on surfaces coated with 4000 HU/mL influenza virus particles and ICAM-1. Each data point on the graph represents the SD of radii for a single cell, and the median ± SE of the median is shown for n = 45-63 cells per donor. NS indicates not significant, analyzed by 1-way ANOVA.