BrdU Incorporation Assay for Cell Proliferation

Verification of synchronization and determination of drug effects was performed by measuring the incorporation of bromo-deoxyuridine (BrdU) into replicating foci within nuclei. At times indicated, cells were pulse-labeled with 15 µM BrdU for 30 min, fixed with 4% paraformaldehyde, and analyzed by standard immunofluorescent techniques^{36 (link), 50 (link)} with an anti-BrdU monoclonal antibody (Roche, clone no. BMC9318, RRID:AB_2313622). The average counts of three fields of 100 or more cells were used to determine the percentages of BrdU-labeled nuclei, +/− 1 s.d.

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Xiang S., Luo X., Welch D., Reed D.R, & Alexandrow M.G. (2023). Identification of Selective ATP-Competitive CMG Helicase Inhibitors for Cancer Intervention that Disrupt CMG-Replisome Function. Research Square.

Publication Preprint 2023

BMC9318

Bromo deoxyuridine Cells Clone Immunofluorescent Monoclonal antibody Nuclei Paraformaldehyde Pulse

Corresponding Organization :

Other organizations : Moffitt Cancer Center

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Variable analysis

independent variables

Exposure of cells to a drug

dependent variables

Incorporation of bromo-deoxyuridine (BrdU) into replicating foci within nuclei
Percentage of BrdU-labeled nuclei

control variables

Pulse-labeling of cells with 15 μM BrdU for 30 min
Fixing cells with 4% paraformaldehyde
Analyzing cells using standard immunofluorescent techniques with an anti-BrdU monoclonal antibody

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

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