Pohl K, Stierhof Y D
Max-Planck-Institut für Biologie, Tübingen, Germany.
Microsc Res Tech. 1998 Jul 1;42(1):59-65. doi: 10.1002/(SICI)1097-0029(19980701)42:1<59::AID-JEMT7>3.0.CO;2-M.
In conventional immunoelectron microscopy (IEM), very small colloidal gold particles (0.8-3 nm), or the gold compound Nanogold (1.4 nm) are silver-enhanced for easy detection. However, silver enhancement has drawbacks. First, the silver layer is dissolved during fixation with osmium tetroxide, even if the concentration and incubation time are strongly reduced during pre-embedding labeling experiments in transmission electron microscopic (TEM) and scanning electron microscopic (SEM) studies. Second, after exposure to the electron beam the silver layer may migrate on the section or the whole particles may disappear. Sometimes silver migration can be observed even without irradiation. This effect strongly hampers reinvestigation of previously inspected areas, after some time of storage. In both cases, gold chloride treatment after silver enhancement is sufficient to completely protect the silver-enhanced 1 nm gold markers. Gold chloride treatment is part of the so-called "gold toning" procedure, which is a method used to substitute and/or cover the silver by a layer of gold. It can be applied in TEM and SEM experiments. As a serious drawback, gold chloride treatment slightly reduces the size of both unenhanced and silver-enhanced gold particles and can lead to disintegrated silver/gold particles. Therefore, this technique is useful for pre-embedding IEM, on-(resin)section, and ultrathin cryosection labeling experiments. However, it appears to be unsuitable for double-labeling studies using different gold sizes, for quantitation experiments, and in SEM.
在传统免疫电子显微镜（IEM）中，非常小的胶体金颗粒（0.8 - 3纳米）或金化合物纳米金（1.4纳米）会进行银增强处理以便于检测。然而，银增强存在缺点。首先，在用四氧化锇固定过程中，银层会溶解，即使在透射电子显微镜（TEM）和扫描电子显微镜（SEM）研究的预包埋标记实验中强烈降低了其浓度和孵育时间。其次，在电子束照射后，银层可能会在切片上迁移，或者整个颗粒可能会消失。有时即使没有照射也能观察到银迁移。在储存一段时间后，这种效应严重妨碍了对先前检查区域的重新研究。在这两种情况下，银增强后进行氯化金处理足以完全保护银增强的1纳米金标记物。氯化金处理是所谓“金调色”程序的一部分，这是一种用一层金替代和/或覆盖银的方法。它可应用于TEM和SEM实验。作为一个严重的缺点，氯化金处理会略微减小未增强和银增强的金颗粒的尺寸，并可能导致银/金颗粒解体。因此，该技术适用于预包埋IEM、（树脂）切片和超薄冷冻切片标记实验。然而，它似乎不适用于使用不同金颗粒尺寸的双重标记研究、定量实验以及SEM。
