Butch E R, Lau H H, Shaw K L, Smolarek T A, Schmerold I, Anderson J N, Baird W M, Yagi H, Jerina D M
Department of Medicinal Chemistry, School of Pharmacy and Pharmacal Sciences, Purdue University, West Lafayette, IN 47907.
Carcinogenesis. 1992 May;13(5):895-9. doi: 10.1093/carcin/13.5.895.
Polyclonal antibodies were developed in New Zealand White rabbits against DNA modified with diastereomeric benzo[c]phenanthrene-3,4-diol-1,2-epoxide (B[c]PhDE)-1 (4-hydroxyl and epoxide cis) and B[c]PhDE-2 (4-hydroxyl and epoxide trans). Antiserum developed against B[c]PhDE-2-DNA was stereoselective. In competitive ELISA assays using wells coated with 160 fmol B[c]PhDE-2-DNA adducts, B[c]PhDE-2-DNA gave 50% inhibition at 200 fmol adducts/well. B[c]PhDE-1-DNA required a 10-fold higher amount of adducts/well to give 50% inhibition. Benzo[a]pyrene-7,8-diol-9,10-epoxide-2-DNA and 7,12-dimethylbenz[a]anthracene-3,4-diol-1,2-epoxide-1-DNA caused only a 30% inhibition even at the highest doses tested (greater than 4000 fmol adducts/well). For antiserum developed against B[c]PhDE-1-DNA, 50% inhibition required 570 fmol B[c]PhDE-1-DNA adducts in wells coated with 100 fmol B[c]PhDE-1-DNA adducts. 7,12-Dimethylbenz[a]anthracene-3,4-diol-1,2-epoxide-1-DNA and B[c]PhDE-2-DNA were also effective competitors: they caused 50% inhibition at 1900 and 1800 fmol adducts/well respectively. In contrast, benzo[a]pyrene-7,8-diol-9,10-epoxide-2-DNA gave no inhibition at the highest dose of competitor tested (4050 fmol adducts/well). Antisera from three rabbits immunized with B[c]PhDE-2-DNA demonstrated similar antigen specificities. The properties of these antisera differ from those reported previously for antibodies developed against benzo[a]pyrene-DNA in that they show selectivity for DNA modified by specific hydrocarbon diolepoxides, in one case for B[c]PhDE-2-DNA and in the other for B[c]PhDE-DNA or 7,12-dimethylbenz[a]anthracene-3,4-diol-1,2-epoxide-1-DNA. The specificity of these antisera will facilitate analysis of the modification of DNA by different polycyclic aromatic hydrocarbon diolepoxides.
针对用非对映体苯并[c]菲-3,4-二醇-1,2-环氧化物（B[c]PhDE）-1（4-羟基和环氧化物顺式）和B[c]PhDE-2（4-羟基和环氧化物反式）修饰的DNA，在新西兰白兔中制备了多克隆抗体。针对B[c]PhDE-2-DNA产生的抗血清具有立体选择性。在使用包被有160 fmol B[c]PhDE-2-DNA加合物的孔进行的竞争性ELISA试验中，B[c]PhDE-2-DNA在200 fmol加合物/孔时产生50%的抑制。B[c]PhDE-1-DNA需要高10倍的加合物量/孔才能产生50%的抑制。苯并[a]芘-7,8-二醇-9,10-环氧化物-二-DNA和7,12-二甲基苯并[a]蒽-3,4-二醇-1,2-环氧化物-1-DNA即使在测试的最高剂量（大于4000 fmol加合物/孔）下也仅产生30%的抑制。对于针对B[c]PhDE-1-DNA产生的抗血清，在包被有100 fmol B[c]PhDE-1-DNA加合物的孔中，50%的抑制需要570 fmol B[c]PhDE-1-DNA加合物。7,12-二甲基苯并[a]蒽-3,4-二醇-1,2-环氧化物-1-DNA和B[c]PhDE-2-DNA也是有效的竞争者：它们分别在1900和1800 fmol加合物/孔时产生50%的抑制。相比之下，苯并[a]芘-7,8-二醇-9,10-环氧化物-2-DNA在测试的最高竞争剂量（4050 fmol加合物/孔）下没有产生抑制。用B[c]PhDE-2-DNA免疫的三只兔子的抗血清表现出相似的抗原特异性。这些抗血清与先前报道的针对苯并[a]芘-DNA产生的抗体的特性不同，因为它们对由特定烃二环氧物修饰的DNA具有选择性，一种情况下对B[c]PhDE-2-DNA有选择性，另一种情况下对B[c]PhDE-DNA或7,12-二甲基苯并[a]蒽-3,4-二醇-1,2-环氧化物-1-DNA有选择性。这些抗血清的特异性将有助于分析不同多环芳烃二环氧物对DNA的修饰。
