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首页> 外文期刊>Mutation research. Genetic toxicology testing >COMPARISON OF AUTORADIOGRAPHY, LIQUID SCINTILLATION COUNTING AND IMMUNOENZYMATIC STAINING OF 5-BROMO-2'-DEOXYURIDINE FOR MEASUREMENT OF UNSCHEDULED DNA SYNTHESIS AND REPLICATIVE DNA SYNTHESIS IN RAT LIVER
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COMPARISON OF AUTORADIOGRAPHY, LIQUID SCINTILLATION COUNTING AND IMMUNOENZYMATIC STAINING OF 5-BROMO-2'-DEOXYURIDINE FOR MEASUREMENT OF UNSCHEDULED DNA SYNTHESIS AND REPLICATIVE DNA SYNTHESIS IN RAT LIVER

机译：5-溴代2'-脱氧尿嘧啶的放射自显影，液体闪烁计数和免疫染色的比较，用于测定大鼠肝脏中计划外的DNA合成和复制性DNA合成

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Different methods for evaluating unscheduled DNA synthesis (UDS) and replicative DNA synthesis (RDS) were studied in hepatocytes of F344 rats exposed in vivo to dimethylnitrosamine (DMN) or CCl4. Hepatocytes were isolated and incubated in Willi ams' medium E supplemented with either [H-3]thymidine for autoradiography or 5-bromo-2'-deoxyuridine for immunoenzymatic staining, In the method of liquid scintillation counting, the cells were incubated with [H-3]thymidine with or without hydroxyurea. The nuclear fraction was isolated and the incorporation of [H-3]thymidine into nuclear DNA was determined by a liquid scintillation counter. DMN at doses of 0.625-5 mg/kg body weight induced UDS of 1.6-37.9 (0 dose; -6.9) net grainsucleus measured by autoradiography and 337-1377 (0 dose; 177) dpm/mu g DNA in the presence of hydroxyurea measured by a liquid scintillation counter. CCl4 at doses of 50-400 mg/kg body weight induced RDS in 1.5-12.1% (0 dose; 0.12%) and 1.8-14.6% (0 dose; 0.16%) of cells with the methods of autoradiography and immunoenzymatic staining, respectively, and of 2991-24256 (0 dose; 324) dpm/mu g DNA in the absence of hydroxyurea with the method of liquid scintillation counting. Similar dose-dependent induction of UDS and RDS was observed with these methods, These results suggest that the methods of liquid scintillation counting and immunoenzymatic staining have almost the same sensitivity for measuring UDS and RDS as that of autoradiography.
机译：在体内暴露于二甲基亚硝胺（DMN）或CCl4的F344大鼠肝细胞中研究了评估计划外DNA合成（UDS）和复制性DNA合成（RDS）的不同方法。分离肝细胞，并在补充有[H-3]胸苷进行放射自显影或在5-溴-2'-脱氧尿苷进行免疫酶染色的威利氏培养基E中孵育。在液体闪烁计数方法中，将细胞与[H]孵育-3]胸苷有或没有羟基脲。分离核级分，并通过液体闪烁计数器确定[H-3]胸苷掺入核DNA中。 DMN剂量为0.625-5 mg / kg体重时，通过放射自显影测得的UDS为1.6-37.9（0剂量; -6.9）净颗粒/核，而337-1377（0剂量; 177）dpm /μgDNA存在液体闪烁计数器测量的羟基脲含量。 50-400 mg / kg体重的CCl4分别通过放射自显影和免疫酶染色方法在1.5-12.1％（0剂量; 0.12％）和1.8-14.6％（0剂量; 0.16％）细胞中诱导RDS。 ，采用液体闪烁计数法测定在没有羟基脲的情况下的2991-24256（0剂量; 324）dpm /μgDNA。用这些方法观察到相似的剂量依赖性诱导UDS和RDS。这些结果表明，液体闪烁计数和免疫酶染色的方法对UDS和RDS的测量灵敏度与放射自显影几乎相同。

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