EdU Imaging Kits (DAB)

Measuring cell proliferation and cell cycle are a fundamental method to assess cell health, determine genotoxicity, and evaluate drug’s pharmacodynamic effect. The common method is measuring DNA synthesis directly. In previous experiments, there are several approaches such as the incorporation of radioactive nucleosides (3H-thymidine) or BrdU. Here, we introduce one new method, click chemistry - CuAAC (Copper-Catalyzed Azide-Alkyne Cycloaddition), and the use of this reaction in direct measurement of S-phase DNA synthesis in cell cycle.

A nucleoside analog of thymidine, EdU (5-ethynyl-2’-deoxyuridine), can be incorporated into DNA strand during DNA synthesis. The alkynyl group of EdU is a biologically inert group that will undergo an extremely selective reaction with dye’s azido via a CuAAC reaction to afford an 1,2,3-triazole product. EdU and Biotin azide possess biologically unique moieties to label DNA of proliferating cells, producing low backgrounds and high detection sensitivities. This CuAAC reaction affords superior regioselectivity and quantitative transformation under extremely mild conditions.

EdU Imaging Kits (DAB) specifically labels the DNA of proliferating cells after biotin azide is ligated to EdU, then adds horseradish peroxidase-labeled streptavidin (HRP-Streptavidin) to bind biotin, developing by DAB chromogen, and finally visualized by microscopy.

Figure 1: The EdU Imaging Kits (DAB) to detect cell proliferation

Figure 2: Detection of cell proliferation using this kit
No staining of cells was observed in the EdU-free group, and brown staining of cells was evident after using EdU, which was inhibited by the addition of Hydroxyurea (a DNA synthesis inhibitor).