Konttinen Y T, Segerberg-Konttinen M, Nordström D, Bergroth V, Scheinin T, Saari H
Fourth Department of Medicine, Helsinki University Central Hospital, Finland.
J Immunol Methods. 1988 May 25;110(1):19-27. doi: 10.1016/0022-1759(88)90078-6.
An immunoperoxidase-autoradiography double labeling method for analysis of lymphocyte activation markers and DNA synthesis is described. For this study expression of MHC locus II coded Ia antigen, interleukin-2 receptor, transferrin receptor and gp 40/80 glycoprotein was analyzed using monoclonal antibodies in avidin-biotin-peroxidase complex staining combined with visualization of [3H]thymidine incorporation by autoradiography. Compared to spontaneous [3H]thymidine incorporation assay information is obtained at single cell level. In contrast to blast indexes calculated from MGG stained preparates, information on the expression of various functional cell surface structures as well as DNA synthesis is also obtained. Double-assay for lymphocyte phenotype and DNA synthesis by flow cytometry might be preferred to light microscopy, but the widespread use of immunoperoxidase staining and autoradiography may make this new kind of approach more easily available. Other advantages worth considering are the possibility of transporting, staining and counterstaining as microscope slides and the permanent nature of the documentation and morphological information obtained. In our experience, this method seems to be useful for studying resting peripheral blood lymphocytes as well as mitogen and antigen induced changes in the lymphocyte activation state.
本文描述了一种用于分析淋巴细胞活化标志物和DNA合成的免疫过氧化物酶-放射自显影双重标记方法。在本研究中，使用单克隆抗体，通过抗生物素蛋白-生物素-过氧化物酶复合物染色，并结合放射自显影观察[3H]胸腺嘧啶核苷掺入情况，分析了主要组织相容性复合体（MHC）II类基因座编码的Ia抗原、白细胞介素-2受体、转铁蛋白受体和gp 40/80糖蛋白的表达。与自发[3H]胸腺嘧啶核苷掺入试验相比，该方法可在单细胞水平获得信息。与根据改良Giemsa染色（MGG）制备物计算的原始细胞指数不同，该方法还可获得关于各种功能性细胞表面结构表达以及DNA合成的信息。虽然通过流式细胞术对淋巴细胞表型和DNA合成进行双重检测可能比光学显微镜检查更具优势，但免疫过氧化物酶染色和放射自显影的广泛应用可能使这种新方法更容易获得。其他值得考虑的优点包括能够像显微镜载玻片一样进行运输、染色和复染，以及所获得的记录和形态学信息的永久性。根据我们的经验，该方法似乎可用于研究静息外周血淋巴细胞以及有丝分裂原和抗原诱导的淋巴细胞活化状态变化。
