Apoptosis

New tool to determine cell divisions

Cell proliferation can occur in response to many stimuli such as cytokine exposure or a variety of other processes. BD has a new product to help researchers study cell proliferation.


BD Biosciences offers BD Horizon™ Violet Proliferation Dye 450 for the detection of cell proliferation with the violet laser, which facilitates the use of larger panels. This allows the determination of more data from limited samples using multicolor flow cytometry.

VPD450 is a nonfluorescent esterified dye. The ester group allows the dye to enter the cell. Once the dye is inside the cell, esterases cleave off the ester group to convert the dye into a fluorescent product and trap it inside the cell. With each replication event the amount of dye in the cell is decreased, leading to a characteristic pattern.

Measurement of Cell Proliferation with BrdU

BD Biosciences carries a series of antibodies and kits designed for the detection of proliferating cells by measurement of bromodeoxyuridine (BrdU), an analog of the DNA precursor thymidine used to measure de novo DNA synthesis.

Tools and Techniques to Study Cell Proliferation

During the S phase of the cell cycle (DNA synthesis) BrdU is incorporated into the newly synthesized DNA and can be readily detected by anti-BrdU specific antibodies. BD antibodies and kits designed for the detection of BrdU are available for both intracellular flow cytometry and immunohistochemistry and include BD Horizon™ V450 and PerCP-Cy™5.5 formats.

In addition to DNA increases, levels of certain proteins also rise as a result of cell proliferation. For example, Ki67 is an antigen that is expressed in the nucleus of dividing cells. However, during the G0 phase of the cell cycle it is not detected. Ki67 can be combined with other proliferation markers such as BrdU and VPD450 for added confidence. These markers can also be combined with cell surface and other types of markers to gain additional information about cell subsets and their signaling pathways.

Cell cycle analysis of a population stained for incorporated BrdU and total DNA levels (7-AAD).
Cell cycle analysis of a population stained for incorporated BrdU and total DNA levels (7-AAD).
CD4+ enriched mouse splenocytes were loaded with 1 µM VPD450 for 10 minutes. Cells were then stimulated with anti-CD3/CD28 and harvested at the indicated times. Approximately 4 to 6 hours prior to harvest, cells were stimulated with PMA/ionomycin in the presence of BD GolgiStop™ protein transport inhibitor. Cells were fixed and permeabilized, stained for IL-2, and analyzed on a BD™ LSR II flow cytometer.
Cell cycle analysis of HeLa cells treated with Aphidicolin

Cell cycle analysis of HeLa cells treated with Aphidicolin (DNA polymerase inhibitor) monitored by BrdU staining. The images were captured on a BD Pathway™ 855 bioimaging system with a 20x objective and merged using BD Attovision™ software.