Establishment and application of the sensitive assay systems for ultraviolet-induced DNA damage.

紫外DNA损伤敏感检测系统的建立及应用

• 批准号: 11470498
• 负责人: NIKAIDO Osamu
• 金额: $ 9.86万
• 依托单位: Kanazawa University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11470498/
• 关键词: DNA damage; cyclobutane pyrimidine dimer (CPD); (6-4) photoproduct; Capillary electrophoresis; Enzyme-linked immunosorbent assay (ELISA); Dewar photoproduct; monoclonal antibody; Dot-blot analysis; シクロブタン型ビリミジン二量体; ヌクレオチド除去修復; シクロブンタン型ピリミジンニ量体 /

We established the quantitative systems to detect various kinds of ultraviolet (UV)-induced DNA damage by using,1) a capillary electrophoresis combined with laser-induced fluorescence detector. The DNA irradiated with various doses of UVC was incubated with the first antibody recognizing DNA damage, followed by incubation with the second antibody (anti-mouse IgG goat antibody) conjugated with fluorescent dye. The DNA damage-first antibody-second antibody-complex was detected in a UVC dose-dependent manner in the rage of 0-1J/m^2.This higher resolution of the capillary electrophoresis for the detection of cyclobutane pyrimidine dimers (CPD) in UV-irradiated DNA made us possible to assess the biological effect of physiologically relevant doses of ultraviolet light in solar light.2) an enzyme-linked immunosorbent assay (ELISA) conjugated with enhancedchemiluminescence system (ECL). We successfully detected CPD in DNA irradiated with UVC doses up to 1J/m^2 by this ECL-ELISA system.

建立了紫外线对DNA损伤的定量检测体系：1）毛细管电泳-激光诱导荧光检测器。将用各种剂量的UVC照射的DNA与识别DNA损伤的第一抗体一起孵育，然后与缀合有荧光染料的第二抗体（抗小鼠IgG山羊抗体）一起孵育。在0- 1 J/m^2范围内，检测到DNA损伤-第一抗体-第二抗体复合物，且呈剂量依赖性。毛细管电泳检测环丁烷嘧啶二聚体（CPD）的高分辨率，使我们有可能评估生理剂量的紫外光在日光下的生物效应。与增强化学发光体系（ECL）偶联。我们成功地用该体系检测了紫外线照射剂量达1 J/m^2的DNA中的CPD。

项目成果

Tanaka, M.: "Effects of photoreactivation of cyclobutane pyrimidine dimers and pyrimidine (6-4) pyrimidone photoproducts on ultraviolet mutagenesis in SOC-induced repair-deficient Escherichia coli."Mutagenesis. 16(1). 1-6 (2001)

Tanaka, M.：“环丁烷嘧啶二聚体和嘧啶 (6-4) 嘧啶酮光产物的光再活化对 SOC 诱导的修复缺陷型大肠杆菌中紫外线诱变的影响。”诱变。

Budiyanto,A.: "Protective effect of topically applied olive oil against photocarcinogenesis following UVB exposure of mice."Carcinogensis,. 21(11). 2085-2090 (2000)

Budiyanto,A.：“局部涂抹橄榄油对小鼠 UVB 暴露后光致癌的保护作用。”致癌作用，。

Ahmed,N.U.: "High levels of 8-hydoroxy-2'-deoxyguanosine appear in normal human epidermis after a single dose of ultraviolet radiation."Brit. J. Dermatol.. 140. 226-231 (1999)

艾哈迈德，N.U.：“单剂量的紫外线辐射后，正常人表皮中会出现高水平的 8-羟基-2-脱氧鸟苷。”

Ishigaki,Y.: "Development and characterization of a DNA solar dosimeter."J.Photochem.Photobiol.B,. 50(2-3). 184-188 (1999)

Ishigaki,Y.：“DNA 太阳剂量计的开发和表征。”J.Photochem.Photobiol.B,。

Horiki,S.: "Protective effects of sunscreening agents on photocarcinogenesis, photoageing, and DNA damage in XPA gene knock out mice."Arch.Delmatol.Res.. 292(10). 511-518 (2000)

Horiki,S.：“防晒剂对 XPA 基因敲除小鼠的光致癌、光老化和 DNA 损伤的保护作用。”Arch.Delmatol.Res.. 292(10)。