Puromycin wash-out leads to fast recovery of CHMP4B levels in MB. (A) HeLa cells were either untreated or treated for 1 h with puromycin. Cells were then washed and incubated with complete media for either 5 min or 10 min. Cells were then fixed and subjected to immunostaining with anti-acetylated-tubulin and anti-CHMP4B antibodies. The asterisks mark the MB and the white dashed square represents the region of the image used for the inset. Scale bars: 10 μm. Inset scale bars: 2 μm. (B and C) Shown data represents CHMP4B intensity/μm² in the intercellular bridge of 60 cells from three separate experiments. B shows distributions derived from individual cells (each dot represents a single cell). Statistical analysis in C was done on the means and SD were derived from the three individual experiments where experimental means were calculated by averaging values from all the cells from each experiment. Each experiment is represented by a different shade of color. Statistical analysis is represented with a P value. (D and E) Shown data represents CHMP4B intensity/μm² in the cell body of 60 cells from three separate experiments. D shows distributions derived from individual cells (each dot represents a single cell). Statistical analysis in E was done on the means and SD were derived from the three individual experiments where experimental means were calculated by averaging values from all the cells from each experiment. Each experiment is represented by a different shade of color. Statistical analysis is represented with a P value. (F and G) HeLa cells that were untreated or treated for 1 h with puromycin. Cells were then washed and incubated with media for an additional 5 or 10 min. Cells were then fixed and stained with anti-acetylated-tubulin antibodies. The percentage of cells in either telophase (F) or just after abscission (G) was then counted. Shown data are means and SD derived from three independent experiments. Statistical analysis is represented with a P value.