Behavior of unpaired kinetochores. (A) MUG in HeLa cells stably expressing CENP-A–GFP followed by combinational differential interference contrast (top) and three-dimensional fluorescence microscopy (bottom). Kinetochores are extremely dynamic, exhibiting rapid poleward and away from pole movements with a mean velocity of 1.8 ± 0.8 μm/min. Anaphase in this cell (56 s) is marked by cessation of kinetochore dynamics, spindle shortening (compare 49 s with 56 s), and outward movement of chromatin (arrowheads). Not all kinetochores congress before anaphase (arrow). (B) Tracking of a single kinetochore (arrowheads) in the cell shown in A. This as well as many other kinetochores exhibits rapid back and forth movement toward and away from the spindle equator. Time is given in hours/minutes. Bar, 2.5 μm.