5 simulations of increasing particle density, each combined with 8 detection efficiencies represented by percentages of particles missing from the detection, were used to test the performance of the tracking algorithm. The results shown are the averages over 6 repetitions of each simulation. (a) Criteria to assess particle density as related to tracking: Average nearest neighbor distance (upper panel, left y-axis) and fraction of particles with nearest neighbors closer than twice their average frame-to-frame displacement (upper panel, right y-axis), evaluated at 0% detection misses; average number of potential assignments per particle (middle panel, left y-axis) and fraction of particles with > 1 potential assignment (middle panel, right y-axis) in the frame-to-frame linking step, evaluated at 0% detection misses; average number of potential assignments per track segment (lower panel, left y-axis) and fraction of track segments with > 1 potential assignment (lower panel, right y-axis) in the gap closing, merging and splitting step, evaluated at 20% detection misses (at 0% misses, there are no gaps to close). (b–d) Percentage of true and false positives in particle linking (b), gap closing (c) and merging and splitting (d) relative to the ground truth (GT). (e) P-value of the Kolmogorov-Smirnov test comparing the measured and GT lifetime distributions. The 0.05 significance threshold is indicated by a dashed line. In (b–e), the conditions similar to the experimental data are highlighted with a dotted oval.