Bromodeoxyuridine Pulse Labelling Assay: A Technique to Measure Cell Proliferation

- Start with a leukemia cell suspension containing the desired cell density. Centrifuge and resuspend the cells in a culture medium containing serum. Serum addition promotes continuous cell growth and cells move across the different cell cycle phases. Dilute the cells in a bromodeoxyuridine, or BrdU, labeling solution and grow them under physiological conditions for the required duration.

Cells in the synthesis or S-phase take up BrdU-- a thymidine analog-- that replaces thymidine during the DNA replication process. Centrifuge the cells to remove excess BrdU and resuspend the cells in a suitable media. Transfer the cells to a multiple well plate. Allow the cells to grow under physiological conditions for different time periods.

At various time periods, cells progress through the remainder of the S, G2, mitosis, and G1 phases and enter into the next cell cycle, allowing for the estimation of cell proliferation. Perform antibody staining to tag BrdU with fluorochrome-antibody and measure cell proliferation using flow cytometry.

Inside the flow cytometer, fluorochromes absorb the laser energy and fluoresce. The number of proliferating cells directly correlates to the fluorescence obse