Fig. 4. Single molecule tracking with stroboscopic illumination.

(A) Timing diagram for stroboscopic illumination. Each laser pulses (1 ms) is synchronized to the CCD frame time, which lasts time T. (B) Displacement histograms for different values of T. The absolute values of displacement along an arbitrary axis were calculated from 2D Gaussian fittings in two successive image frames. The displacement distribution of nonspecifically bound TFs broadens with time (left), whereas the distribution before induction (right) remains peaked at <100 nm. The contrast between them illustrates the change in TF mobility before and after induction. (C) Mean-square displacement for nonspecifically bound TFs at different time intervals. The red line shows a linear fit of the MSDs. Error bars are calculated as described in the SOM. The fitting agrees well with a normal diffusion in the imaging plane, <Δx²>=4D_efft, with D_eff=0.4 µm²s⁻¹.