Immunohistological examination of the inter- and intracellular distribution of O6-alkylguanine DNA-alkyltransferase in human liver and melanoma.
Immunohistological examination of the inter- and intracellular distribution of O6-alkylguanine DNA-alkyltransferase in human liver and melanoma.
复制标题
O6-烷基鸟嘌呤DNA-烷基转移酶在人肝和黑色素瘤中的细胞内和细胞内分布的免疫组织学检查。
DOI：
10.1038/bjc.1992.270
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发表时间：
1992-08
影响因子：
8.8
通讯作者：
Perinat-Frey, T
中科院分区：
文献类型：
--
作者：
Lee, S M;Rafferty, J A;Elder, R H;Fan, C Y;Bromley, M;Harris, M;Thatcher, N;Potter, P M;Altermatt, H J;Perinat-Frey, T
DNA修复蛋白O6-烷基鸟嘌呤-DNA-烷基转移酶(ATase)的组织和细胞分布是一个重要的问题，它关系到肿瘤和正常组织对甲基三氮烯和亚硝脲等烷化药物化疗方案的反应。为了通过免疫染色来检测这一问题，我们将兔抗血清提纯为明显纯的重组人酶。该抗血清具有高度的特异性和敏感性，在表达ATase的人淋巴母细胞、肝脏和黑色素瘤的粗提物的Western印迹上检测到一条24 kDa的条带。将丙酮或福尔马林固定的人正常肝脏和皮下恶性黑色素瘤的相邻切片与前免疫血清或抗血清反应，用银增强免疫过氧化物酶检测系统定位一抗与抗原的结合。在与免疫前血清或抗原预吸附抗血清反应的切片上，仅见微弱的胞浆，核很少或未见染色。相比之下，使用抗血清，阳性染色细胞中的反应非常强烈，以核为主。在肝脏中，反应的细胞分布存在个体差异，大多数标本中所有可辨别的细胞类型都有染色，而其他标本仅限于肝细胞和胆管上皮细胞。在黑色素瘤切片中，所有可识别的细胞类型都以核染色为主：不同组织样本的染色强度不同，有证据表明有一系列中等染色强度的证据，一些黑色素瘤细胞没有检测到反应。
The tissue and cellular distribution of the DNA repair protein O6-alkylguanine-DNA-alkyltransferase (ATase) is an important question in relation to the response of tumour and normal tissues to chemotherapeutic regimes employing alkylating agents such as methyltriazenes and nitrosoureas. In order to examine this issue by immunostaining, we have raised a rabbit antiserum to apparently pure recombinant human enzyme. The antiserum is highly specific and sensitive, detecting a band at 24 kDa on western blots of crude extracts of ATase-expressing human lymphoblastoid cells, liver and melanoma. Adjacent sections of acetone or formalin fixed normal human liver and subcutaneous malignant melanoma were reacted with preimmune serum or antiserum and an immunoperoxidase detection system with silver enhancement was used to locate binding of the primary antibody to the antigen. In sections reacted with preimmune serum or with antigen-preadsorbed antiserum, only faint cytoplasmic and little or no nuclear staining was seen. In contrast, using antiserum, the reaction in positively staining cells was very intense and predominantly nuclear. In the liver, there was interindividual variation in the cellular distribution of reaction with staining present in all discernable cell types in most samples but confined to the hepatocytes and bile duct epithelial cells in others. In the melanoma sections, all discernable cell types showed mainly nuclear staining: the intensity of staining varied between tissue samples and there was evidence of a range of intermediate staining intensities with some melanoma cells showing no detectable reaction.