The thymidine analog 5′-bromo-2′-deoxyuridine (BrdU) is extensively referenced in the scientific literature as a tool for determining the rates of cell proliferation, as dividing cells incorporate BrdU into newly synthesized DNA during the S phase of the cell cycle. Anti-BrdU antibodies, such as the Mouse Anti-BrdU Antibody, clone Bu20a (MCA2483), are subsequently used to detect and visualize the incorporated BrdU. This is commonly done by immunodetection techniques such as flow cytometry, immunocytochemistry, and immunohistochemistry.
BrdU is a universal tool that has been successfully used for studies in evolutionary diverse organisms ranging from plants, fish, and invertebrates (such as the flatworm Macrostomum lignano) to vertebrates, including mammalian species (Caronia et al. 2010, Huang et al. 2013, Nagar et al. 2002, Verdoodt et al. 2012).
Although BrdU has been successfully established as a tool in many biological systems, the use of BrdU and other thymidine analogs has its limitations. For example, BrdU cannot be readily incorporated into yeast DNA as yeasts lack thymidine nucleoside transporters and thymidine kinases, which are essential for the uptake and incorporation of exogenous thymidine (or its analogs) into newly synthesized DNA (Lengronne et al. 2001). To compensate for the absence of these critical proteins, Saccharomyces cerevisiae strains have been genetically engineered to express exogenous genes, such as the Herpes simplex virus thymidine kinase gene and human equilibrative nucleoside transporters (Lengronne et al. 2001, Viggiani and Aparico 2006).
Due to the structure of DNA, incorporated BrdU is often inaccessible to anti-BrdU antibodies. Therefore, for BrdU detection, it is necessary to denature or cleave the DNA prior to incubation with anti-BrdU antibodies. Commonly used DNA denaturation methods include treatment with hydrochloric acid (HCl) or sodium hydroxide (NaOH) (Ligasová et al. 2017). However, due to the strength of the treatment with acids such as HCl, protein denaturation may occur, which could impair immunostainings (Tkatchenko 2006). Alternative milder treatment options include exposure to nucleases (such as DNase I) or monovalent copper ions (Liboska et al. 2012). Our Mouse Anti-BrdU Antibody, clone Bu20a, has been cited in 13 publications and is supported by flow cytometry and immunocytochemistry protocols.
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