ASSAY OF EXCISED OXIDATIVE DNA LESIONS - ISOLATION OF 8-OXOGUANINE AND ITS NUCLEOSIDE DERIVATIVES FROM BIOLOGICAL-FLUIDS WITH A MONOCLONAL-ANTIBODY COLUMN
ASSAY OF EXCISED OXIDATIVE DNA LESIONS - ISOLATION OF 8-OXOGUANINE AND ITS NUCLEOSIDE DERIVATIVES FROM BIOLOGICAL-FLUIDS WITH A MONOCLONAL-ANTIBODY COLUMN
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DOI：
10.1073/pnas.89.8.3375
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发表时间：
1992-04-15
影响因子：
11.1
通讯作者：
AMES, BN
中科院分区：
文献类型：
--
作者：
PARK, EM;SHIGENAGA, MK;AMES, BN
描述了一种免疫亲和柱，其有助于分析尿液、血浆和从大肠杆菌培养物中分离的培养基中DNA和RNA的氧化损伤产物。在完整的动物中，从DNA切除的病变（加合物）通过循环从细胞中转运并经尿液排泄。在细菌中，DNA加合物直接分泌到培养基中。在任一种情况下，加合物可以作为对DNA的氧化损伤的量度进行测定。已分离出识别8-氧代-7，8-二氢-2 '-脱氧鸟苷（oxo 8dG; 8-羟基-2'-脱氧鸟苷）（DNA氧化损伤的生物标志物）的单克隆抗体，并表征了其底物结合特性。该单克隆抗体对oxo 8dG、未修饰的核苷或Gua衍生物的相对结合亲和力使其适合于制备免疫亲和柱，所述免疫亲和柱极大地促进从各种生物流体中分离oxo 8dG、8-氧代-7，8-二氢鸟嘌呤和8-氧代-7，8-二氢鸟苷。定量分析这些加合物在大鼠尿中喂食无核酸饮食和在培养基中从培养的大肠杆菌。coli的研究表明，无论是在真核生物还是原核生物中，ox 8dG都是ox 8dG的主要修复产物。结果支持我们之前的估计，即平均每天在大鼠细胞中形成和切除约10（5）个DNA氧化损伤。
An immunoaffinity column is described that facilitates the analysis of oxidative damage products of DNA and RNA in urine, blood plasma, and medium isolated from cultures of Escherichia coli. In intact animals, lesions (adducts) excised from DNA are transported from the cell through the circulation and excreted in urine. In bacteria, DNA adducts are excreted directly into the medium. In either case, the adducts can be assayed as a measure of oxidative damage to DNA. A monoclonal antibody that recognizes 8-oxo-7,8-dihydro-2'-deoxyguanosine (oxo8dG; 8-hydroxy-2'-deoxyguanosine), a biomarker of oxidative damage to DNA, has been isolated, and its substrate binding properties have been characterized. The relative binding affinities of this monoclonal antibody for oxo8dG, unmodified nucleosides, or derivatives of Gua made it suitable for the preparation of immunoaffinity columns that greatly facilitate the isolation of oxo8dG, 8-oxo-7,8-dihydroguanine, and 8-oxo-7,8-dihydroguanosine from various biological fluids. Quantitative analysis of these adducts in urine of rats fed a nucleic acid-free diet and in the medium from cultures of E. coli suggests that oxo8-7,8-dihydroguanine is the principal repair product from ox8dG in DNA of both eukaryotes and prokaryotes. The results support our previous estimate of about 10(5) oxidative lesions to DNA being formed and excised in an average rat cell per day.