Decreased motility by WASp-null BMDCs in response to CCL21. (A) Control and WASp-null DCs were exposed to 100 ng/mL CCL21, and their migration was followed by time-lapse microscopy for 5 hours. Data on cell tracks were collected using Openlab software and further analyzed using a Mathematica notebook to generate data on cell trajectories and cell speed. In the absence of chemokine (CM), little persistent movement was observed in either control (top row) or WASp-null (bottom row) LPS-matured DCs (mean cell speed = 0.073 ± 0.0076 μm/sec and 0.072 ± 0.013 μm/sec, respectively; mean distance traveled = 1314 ± 137 μm and 858 ± 97 μm, respectively; not significant, P > .05). Stimulation with CCL21 (CM + CCL21) dramatically increased both the speed of movement and the persistence of directional migration in control cells, whereas WASp-null DCs failed to migrate effectively (mean cell speed = 0.119 ± 0.0082 μm/sec and 0.076 ± 0.013 μm/sec, respectively; mean distance traveled 2143 ± 145 μm and 1031 ± 110 μm, P < .01). Occasional poorly adhered cells, which moved out of the frame, were excluded from the analysis. The cell tracks from 1 of 2 separate experiments were merged into a single file for analysis, giving a total of 22 to 25 cells per treatment. The x- and y-axes are scaled in micrometers (μm). Statistical differences in migration were calculated using a Student t test. (See also supplemental movie 2A-D). (B) Levels of CCR7 expression in matured cells from control (left) and WASp-deficient (right) animals were equivalent when measured by flow cytometry. Unfilled histograms represent an isotype control. Filled histograms indicate specific expression.