Fig. 2. 3F3/2 immunostaining on mitotic human primary fibroblasts (MRC-5 cells). (A) MRC-5 cell arrested in mitosis by nocodazole treatment. All the kinetochores show 3F3/2 staining caused by the presence of the phosphoepitope recognized by the 3F3/2 antibody on unattached kinetochores. (B) Metaphase cell after release from a nocodazole-induced mitotic block. The 3F3/2 antibody does not stain kinetochores, which are aligned at the metaphase plate and under tension. The 3F3/2 antibody does stain spindle poles (arrows). (C,D) Anaphase cells with lagging chromosomes (arrows) after release from the mitotic block. There is no 3F3/2 staining on kinetochores of lagging chromosomes, nor on kinetochores of chromosomes correctly segregated to the spindle poles. DNA is pseudocolored in red; 3F3/2 antibody is shown in yellow. Bar, 5 µm.