Bendayan M
J Histochem Cytochem. 1981 Apr;29(4):531-41. doi: 10.1177/29.4.6265546.
A cytochemical technique for the ultrastructural localization of substrates using enzyme-gold complexes is reported. RNase A and DNase I have been labeled with gold particles. The RNase-gold and dNase-gold complexes obtained were applied on thin sections of glutaraldehyde-fixed and Epon-embedded tissues. Different cellular compartments were labeled by these enzyme-gold complexes. Using the RNase-gold complex the rough endoplasmic reticulum appeared decorated with gold particles. The gold marker was also present over the nucleus, especially over the nucleolus; mitochondria were weakly labeled. Using the DNase-gold complex, gold particles were concentrated over the euchromatin of the nucleus and the mitochondria. The heterochromatin and the nucleolus showed a less intense labeling. For both enzyme-gold complexes, the Golgi area, the secretory granules and the extracellular space appeared free of label. In those control conditions where the substrates were added to the enzyme-gold complexes a major reduction in the labeling was observed. A quantitative evaluation of the labeling was performed. This evaluation confirmed the qualitative observations and the marked reduction of labeling occurring under the control conditions. The combination of the specificity of the enzyme-substrate interactions with the size and electron density of the gold particles and the good ultrastructural preservation of the tissues resulted in a very specific labeling with high resolution. These results demonstrate the possibility of detecting substrates by means of enzyme-gold complexes at the electron microscope level.
报道了一种使用酶-金复合物对底物进行超微结构定位的细胞化学技术。核糖核酸酶A和脱氧核糖核酸酶I已用金颗粒标记。将得到的核糖核酸酶-金和脱氧核糖核酸酶-金复合物应用于经戊二醛固定和环氧树脂包埋的组织薄片上。这些酶-金复合物标记了不同的细胞区室。使用核糖核酸酶-金复合物时，粗面内质网表面出现金颗粒标记。细胞核尤其是核仁上也有金标记；线粒体标记较弱。使用脱氧核糖核酸酶-金复合物时，金颗粒集中在细胞核的常染色质和线粒体上。异染色质和核仁的标记较弱。对于这两种酶-金复合物，高尔基体区域、分泌颗粒和细胞外空间均未出现标记。在将底物添加到酶-金复合物的对照条件下，观察到标记显著减少。对标记进行了定量评估。该评估证实了定性观察结果以及对照条件下标记的显著减少。酶-底物相互作用的特异性与金颗粒的大小和电子密度以及组织良好的超微结构保存相结合，产生了具有高分辨率的非常特异性的标记。这些结果证明了在电子显微镜水平上借助酶-金复合物检测底物的可能性。
