Yang X Y, DeLeo V, Santella R M
Cancer Res. 1987 May 1;47(9):2451-5.
Monoclonal antibodies specific for DNA damaged by 8-methoxypsoralen (8-MOP) plus ultraviolet A (UVA) light were used to study adduct formation in human keratinocytes and mouse and rat skin in vivo. This antibody does not cross-react with nonmodified DNA or free 8-MOP. Sensitive competitive enzyme-linked immunosorbent assays with color or fluorescence endpoints were used to quantitate adducts on DNA isolated from treated keratinocytes or skin samples. Localization of 8-MOP-DNA adducts was studied by indirect immunofluorescence with fluorescein-conjugated anti-mouse-IgG antibodies. When cultured keratinocytes were treated with 8-MOP and UVA, immunofluorescence was localized in the nucleus. There was no fluorescence in untreated control cells or treated cells incubated with nonspecific serum. Comparison of intensity of immunofluorescence staining with quantitation of adduct levels by enzyme-linked immunosorbent assay indicated that the limit of sensitivity of the immunofluorescence technique is 9.0 fmol adduct/micrograms DNA or 2.9 adducts/10(6) nucleotides.
使用对8-甲氧基补骨脂素（8-MOP）加紫外线A（UVA）光损伤的DNA具有特异性的单克隆抗体，在体内研究人角质形成细胞以及小鼠和大鼠皮肤中的加合物形成情况。该抗体与未修饰的DNA或游离的8-MOP无交叉反应。采用具有颜色或荧光终点的灵敏竞争性酶联免疫吸附测定法，对从经处理的角质形成细胞或皮肤样本中分离出的DNA上的加合物进行定量。通过用异硫氰酸荧光素偶联的抗小鼠IgG抗体进行间接免疫荧光，研究8-MOP-DNA加合物的定位。当培养的角质形成细胞用8-MOP和UVA处理时，免疫荧光定位于细胞核。未处理的对照细胞或用非特异性血清孵育的处理细胞中无荧光。免疫荧光染色强度与通过酶联免疫吸附测定法定量加合物水平的比较表明，免疫荧光技术的灵敏度极限为9.0 fmol加合物/微克DNA或2.9个加合物/10⁶个核苷酸。
