DNAM-1^posNK cell subsets display more dynamic and deliberate migration behavior than CD56^dimDNAM-1^negNK cells. CD56^brightDNAM-1^pos, CD56^dimDNAM-1^pos, and CD56^dimDNAM-1^neg NK cell subsets were fluorescence-activated cell sorter purified from healthy donor PBMCs. Cells were stimulated overnight with IL-12 (10 ng/mL), IL-15 (100 ng/mL), and IL-18 (50 ng/mL). Time-lapse microscopy was performed by using an Olympus Xcellence IX81 microscope controlled by Xcellence RT software. Images were taken every 40 seconds for 40 to 90 minutes. Images were collected by using a 20× LUCPlanFLN lens with 0.45 NA and Olympus F-View II camera. (A) Representative images. (B) Single-cell tracking map of NK cell trajectories. Mean velocity (C), length of tracks (D), and displacement from origin (E) of individual NK cells were calculated. Each data point represents 1 single randomly selected NK cell tracked over 40 minutes. Results are displayed as the mean ± SEM of at least 6 representative time-lapse videos obtained from 3 independent experiments. Data were analyzed by using a 1-way ANOVA followed by a Tukey multiple comparisons post hoc test. ***P < .001; ****P < .0001.