Video 1. Related to Figure 1.
Illustration of defocalization bias. Illustration of a single-particle tracking experiment with two subpopulations (one ‘immobile’, D = 0.001 µm²/s, the other ‘free’, D = 4 µm²/s with a 1:1 ratio, observed using 20 ms time interval). The red region corresponds to the axial detection range (1 µm) and molecules randomly appear when they photo-activate. For each trajectory, the detected localizations inside the detection range are shown as red spheres and undetected localizations outside the detection range are shown as white spheres. Each particle has a mean lifetime of 15 frames, 25 nm localization error and trajectories consisting of at least two frames are plotted. Epi illumination is assumed. The SPT data was simulated and plotted using simSPT (available at https://gitlab.com/tjian-darzacq-lab/simSPT).