Fascin decreases the rate of cofilin cluster initiation on single actin filaments. (A–C) Two side-by-side populations of single actin filaments are exposed in a microfluidics chamber to 200 nM cofilin, 0.15 µM actin, and either no or (A) 200 nM, (B) 500 nM, (C) 1 µM fascin (n = 54, 58, 60 segments with fascin, and n = 57, 53, 60 segments without fascin). Fraction of 5-µm segments of single actin filaments with at least one cofilin cluster, as a function of time. 95% confidence intervals are shown as shaded surfaces. Curves are fitted with a single exponential function to derive the cofilin cluster initiation rate for each population. (D) Cofilin cluster initiation rate fold difference between the population of single actin filaments exposed to fascin or not. Error bars for each condition are 95% confidence intervals, based on the sample sizes of the two survival fractions.