Figure 11.

Velocity at small fence sizes. Average velocity (lines with symbols), individual maximum velocity (*), and minimum velocity (×) measurements are shown for objects associated with tracks of small fence sizes from three experiments (see legend). Average velocity was calculated as the mean of all velocity measurements from objects meeting cell specific criteria for area with tracks consisting of 3 or more segments, pooled according to fence size and experiment. Consistency between experiments reflects consistency in the rate at which stage noise and object segmentation variability are exhibited. By limiting fence size, we also limit the maximum possible displacement, and we would expect this to be reflected in the maximum observable velocity. It is. Consider tracks with a fence size of 20 microns (10 pixels); this is near the maximum observable displacement for these objects between any two time-points*. Since the images were taken at 30 minute intervals, the corresponding maximum permissible velocity is 20/30 um/min, or 0.67 um/min, which corresponds to data from all three experiments at a fence size of 20 um. These data, in addition to manually constructed images (see Methods), confirm soundness within the system for accurate velocity measurement. (*Fence size is not strictly the maximum observable displacement. A slightly more sophisticated definition of fence size would use the diagonal from maximum x and y values, which would result in slightly greater fence sizes for objects displaced in random directions.)