[关键词]
[摘要]
目的 建立大鼠肝细胞和外周血淋巴细胞的体内彗星试验方法,并应用该方法对遗传毒性阳性物甲磺酸乙酯(EMS)和环磷酰胺(CPA)进行检测。方法 选择SD雄性大鼠为实验动物,分别给予不同剂量EMS和CPA。给药结束后,采集外周血样本和摘取动物左侧肝叶组织样本,铺制成单细胞凝胶玻片。玻片经裂解、解旋、电泳、中和、脱水等步骤后,得到可观察的实验标本。染色标本,并在40×荧光显微镜下观察拍照,使用专业分析软件对单细胞电泳图像进行分析和测定。结果 成功建立了基于大鼠肝细胞和外周血淋巴细胞的体内彗星试验方法。经该方法检测,EMS结果为阳性,CPA结果为阴性,EMS适合作为本方法的阳性对照。结论 成功建立大鼠体内彗星试验方法,并可应用于遗传毒性检测。
[Key word]
[Abstract]
Objective To establish the comet assay in vivo method used for rat liver and peripheral blood lymphocyte cells and detect the genotoxicity of ethyl methanesulfonate (EMS) and cyclophosphamide (CPA) Methods Sprague-Dawley (SD) male rats were treated with series doses of EMS and CPA. Peripheral blood sample and tissue in left lobe of liver were collected to prepare single cell gel suspension and pave the slides. The slides were processed with complex steps containing lysis, unwinding, electrophoresis, neutralizing, and dehydrating to get the scoring specimen. To dye the specimens, observe the comet cells under fluorescence microscope (40×), and then photograph. The cell images were analyzed with special software and determined by electrophoresis. Results EMS showed positive reaction while CPA was negative in two tissues. It was convinced that EMS was appropriate as positive reagent in this assay. Conclusion The method of rat comet assay in vivo is successfully established and it could be used to genotoxicity detection.
[中图分类号]
[基金项目]
重大新药创制科技重大专项(2008ZX09305-002)