Otsuki Y, Maxwell L E, Magari S, Kubo H
Department of Anatomy, Osaka Medical College, Japan.
J Histochem Cytochem. 1990 Aug;38(8):1215-21. doi: 10.1177/38.8.2365991.
We used the immunogold-silver staining method (IGSS) for detection of lymphocyte cell surface antigens with monoclonal antibodies in light and electron microscopy and compared this procedure with the immunogold staining method. Two different sizes of colloidal gold particles (5 nm and 15 nm) were used in this study. Immunolabeling on cell surfaces was visualized as fine granules only by IGSS in light microscopy. The labeling density (silver-gold complexes/cell) and diameters of silver-enhanced gold particles on cell surfaces were examined by electron microscopy. Labeling density was influenced not by the enhancement time of the physical developer but by the size of the gold particles. However, the development of shells of silver-enhanced gold particles correlated with the enhancement time of the physical developer rather than the size of the colloidal gold particles. Five-nm gold particles enhanced with the physical developer for 3 min were considered optimal for this IGSS method because of reduced background staining and high specific staining in the cell suspensions in sheep lymph. Moreover, this method may make it possible to show the ultrastructure of identical positive cells detected in 1-micron sections counterstained with toluidine blue by electron microscopy, in addition to the percentage of positive cells by light microscopy.
我们采用免疫金银染色法（IGSS），在光学显微镜和电子显微镜下用单克隆抗体检测淋巴细胞细胞表面抗原，并将该方法与免疫金染色法进行比较。本研究使用了两种不同大小的胶体金颗粒（5纳米和15纳米）。在光学显微镜下，仅通过免疫金银染色法可将细胞表面的免疫标记物可视化为细颗粒。通过电子显微镜检查细胞表面银增强金颗粒的标记密度（银-金复合物/细胞）和直径。标记密度不是受物理显影剂增强时间的影响，而是受金颗粒大小的影响。然而，银增强金颗粒壳的显影与物理显影剂的增强时间相关，而不是与胶体金颗粒的大小相关。由于在绵羊淋巴的细胞悬液中背景染色减少且特异性染色高，用物理显影剂增强3分钟的5纳米金颗粒被认为是该免疫金银染色法的最佳选择。此外，除了通过光学显微镜观察阳性细胞的百分比外，该方法还可能使通过电子显微镜显示在经甲苯胺蓝复染的1微米切片中检测到的相同阳性细胞的超微结构成为可能。
