Spatiotemporal dynamics of tractions during neutrophil chemotaxis. (A) dHL-60 cells were allowed to migrate toward chemoattractant-containing micropipette (fMLP, 10μM) on a FN-coated elastic polyacrylamide gel for the times indicated. The speed for cell migration is approximately 2.0 μm/minute, lower than the speed of cells on FN-coated glass (∼ 2.4 μm/minute). The cells also exhibit normal tail retraction on the elastic gel (data not shown). Traction maps of the cell are shown. Pseudocolor bar representing tractions is given in Pascal (Pa). Scale bar represents 5 μm. Arrow indicates the direction of fMLP gradient. The leading edge of a polarized neutrophil was defined as the area within the first 3 μm of the cell (marked by a white line), whereas the rest of the cell was defined as the trailing edge (“Polyacrylamide gel substrates, TFM, and data analysis”). The image series shows part (7.2 seconds, for which the cell traveled ∼ 0.24 μm) of the whole migratory response. The video of the cell in panel A is available in supplemental data. (B) Time series of traction maps from panel A (with 3 additional time points) was analyzed by a customized MATLAB program to determine the average tractions in both leading edge (front) and trailing edge (back) of the cells in a time-dependent manner. The graph shows part (∼ 9 seconds) of the whole migratory response. The x-axis indicates time in seconds; y-axis is in Pascal (Pa). The mean levels of tractions at the leading and the trailing edges were comparable. A graph of another cell with a longer migratory response is shown in supplemental Figure 1A. (C) PSD plots of tractions at the leading (left panel) and the trailing edge (right panel) of a migratory dHL-60 cell. PSD plots were generated based on the results from Fourier analysis of the traction values. The y-axis represents the power spectral density normalized to the highest peak value (= 1); x-axis shows the oscillation frequency (Hertz; top) or period (seconds; bottom). Nine cells were analyzed, and a representative cell is shown. PSD plots of tractions in 3 cells combined are shown in supplemental Figure 1B. (D) PSD plots of tractions at the leading edge (left panel) and the trailing edge (right panel) of a migratory primary neutrophil. Six cells were analyzed, and data from a representative cell are shown. (E) Left panel: Cross-correlation between tractions at the leading edge and the trailing edge against time offset during migration for individual dHL-60 cells. Dotted lines indicate zero offset. Note that the back traction lags the front traction as indicated by the maximum cross-correlation at time offset of 0.8 seconds. Data from 3 representative cells are shown. Time bar represents 24 seconds. Right panel: Summary of time offsets between leading edges and trailing edges (n = 9 cells) in dHL-60 cells. (F) Left panel: Cross-correlation between tractions at the leading and the trailing edges against time offset during migration for individual primary neutrophils. Data from 2 representative cells are shown. Right panel: Summary of time offsets between leading edges and trailing edges (n = 6 cells) in primary neutrophils.