Evaluation of 5-ethynyl-2′-deoxyuridine staining as a sensitive and reliable method for studying cell proliferation in the adult nervous system

Chenbo Zeng, Fenghui Pan, Lynne A. Jones, Miranda M. Lim, Elizabeth A. Griffin, Yvette I. Sheline, Mark A. Mintun, David M. Holtzman, Robert H. Mach

Research output: Contribution to journalArticlepeer-review

159 Scopus citations


Recently, a novel method for detection of DNA synthesis has been developed based on the incorporation of 5-ethynyl-2′-deoxyuridine (EdU), a thymidine analogue, into cellular DNA and the subsequent reaction of EdU with a fluorescent azide in a copper-catalyzed [3 + 2] cycloaddition ("Click" reaction). In the present study, we evaluated this method for studying cell proliferation in the adult central nervous system in comparison with the "gold standard" method of 5-bromo-2′-deoxyuridine (BrdU) staining using two behavioral paradigms, voluntary exercise and restraint stress. Our data demonstrate that the number of EdU-positive cells in the dentate gyrus of the hippocampus (DG) slightly increased in an EdU dose-dependent manner in both the control and voluntary exercise (running) mouse groups. The number of EdU-labeled cells was comparable to the number of BrdU-labeled cells in both the control and running mice. Furthermore, EdU and BrdU co-localized to the same cells within the DG. Voluntary exercise significantly increased the number of EdU- and BrdU-positive cells in the DG. In contrast, restraint stress significantly decreased the number of EdU-positive cells. The EdU-positive cells differentiated into mature neurons. EdU staining is compatible with immunohistochemical staining of other antigens. Moreover, our data demonstrated EdU staining can be combined with BrdU staining, providing a valuable tool of double labeling DNA synthesis, e.g., for tracking the two populations of neurons generated at different time points. In conclusion, our results suggest that EdU staining is a fast, sensitive and reproducible method to study cell proliferation in the central nervous system.

Original languageEnglish (US)
Pages (from-to)21-32
Number of pages12
JournalBrain research
StatePublished - Mar 10 2010
Externally publishedYes


  • 5-bromo-2′-deoxyuridine
  • 5-ethynyl-2′-deoxyuridine
  • BrdU
  • adult neurogenesis
  • hippocampus

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Clinical Neurology
  • Developmental Biology


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