Incorporation of EdU or 5-ethynyl-2′-deoxyuridine allows the sensitive detection of DNA synthesis in cells using “click chemistry."
In 2008, Salic and Mitchinson reported the EdU based method for a fast and sensitive detection of DNA synthesis in vivo. No sample fixation or DNA denaturation is required and biological structures are preserved as well.
As reported, the process works in cultured cells, the intestine, and brain of whole animals in which the cells incorporate 5-ethynyl-2'-deoxyuridine (EdU) into replicating DNA. After incorporation, the terminal alkyne group is available for the reaction with organic azides using Cu(I)-catalyzed click reactions.
Figure 1: Schematics of the copper (I) catalyzed click chemistry.
The availability of the 5-Ethynyl-dU-CE Phosphoramidite also allows incorporation of EdU into DNA during solid phase synthesis.
Salic and Mitchinson found that EdU incorporated into the DNA of proliferating mammalian cells, for example, into DNA replicated in vitro in Xenopus egg extracts by addition to the cycling extract preparations. Varying ascorbic acid and fluorescent azide concentrations allow controlling the amount of the fluorescent signal generated during EdU detection.
Salic and Mitchinson also indicate that the EdU-labeling method is well suited for optical imaging of cellular DNA at a nanometer resolution.
Salic, Adrian & Mitchison, Timothy J.; A chemical method for fast and sensitive detection of DNA synthesis in vivo. Proc Natl Acad Sci USA 2008 105 (7) 2415 – 2420. http://www.pnas.org/content/105/7/2415.abstract