Measurement of DNA damage by terminal deoxynucleotidyl transferase reaction

Masaaki Kurasaki, Yongkun Sun, Miyako Komori, Miki Miyajima, Toshiyuki Hosokawa, Takeshi Saito

Environmental Adaptation Science, Division of Environmental Science Development, Graduate School of Environmental Science, Hokkaido University, Sapporo, Japan.
Group of Environmental Adaptation Science, Faculty of Environmental Earth Science, Hokkaido University, Sapporo, Japan.
Higher Education Research and Development, Division Institute for the Advancement of Higher Education, Hokkaido University, Sapporo, Japan.
Laboratory of Environmental Health Sciences, Faculty of Health Sciences, Hokkaido University, Sapporo, Japan.
Laboratory of Environmental Health Sciences, Graduate School of Health Science, Hokkaido University, Sapporo, Japan.

DOI: 10.4236/abc.2012.23030                

Abstract

An improved terminal deoxynucleotidyl transferase- mediated dUTP-biotin nick end labeling method for the quantification of DNA damage in tissues and cultured cells was developed. Many reports have revealed that histochemistry of DNA damage can be visualized using immunohistochemistry for the terminal deoxynucleotidyl transferase reaction in tissue sections. However, few reports have described quantification of DNA damage in tissues or cells. In this study, to estimate the degree of DNA damage, the confirmed method for histochemistry using biotinylated dUTP and deoxynucleotidyl transferase was applied to label the cleaved DNA ends caused by DNA damage in tissues or cells. After end-labeling, avidin-conjugated peroxidase was reacted. A significant correlation was observed between numbers of cleaved DNA ends and peroxidase activity after the reaction. The obtained signals for presented method showed higher than those for ordinary method, and correlate with degree of DNA damage caused by serum deprivation and chemical dose. In addition, DNA damage caused by apoptosis in cells treated with 6-hydroxydopamine or Cu and in the tissues of rats administered a diet containing no Zn could be evaluated quantitatively using the present method.

Keywords

DNA Damage; Quantification; TUNEL; Apoptosis; PC12 Cells

Conflicts of Interest

The authors declare no conflicts of interest.

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