Eskelinen S, Peura R
Department of Physiology, University of Oulu, Finland.
Scanning Microsc. 1988 Sep;2(3):1765-74.
The use of colloidal gold particles for locating cell surface components by scanning electron microscopy (SEM) has been restricted due to difficulties in the identification of these gold particles under SEM. It is shown here how the gold particles bound to cell surfaces can be located and identified under SEM using the secondary electron imaging (SEI) mode with an energy dispersive X-ray microanalyzer (EDS). This enables reliable identification of gold particles and good quality micrographs of the cells to be achieved at the same time. The distribution of receptors for two lectins, concanavalin A (ConA) and wheat germ agglutinin (WGA), on the surface of cultured Raji cells and human erythrocytes is presented as an example. Raji cells and erythrocytes were fixed with glutaraldehyde, post-fixed with a glutaraldehyde-tannic acid mixture and then incubated with ConA- or WGA-coated gold particles. After dehydration and critical point drying, the specimen filters were mounted on copper stubs and coated with carbon. The cells were examined on a JEOL TEMSCAN 100CX II electron microscope. The gold particles could be identified with the EDS analyzer, which was able to detect the Au spectrum when the electron beam was focused on single gold particles using a magnification of 100,000 or more. High-resolution photographs of the same cells were obtained up to the same magnification of 100,000.
由于在扫描电子显微镜（SEM）下难以识别胶体金颗粒，因此通过扫描电子显微镜定位细胞表面成分时，胶体金颗粒的使用受到了限制。本文展示了如何在带有能量色散X射线微量分析仪（EDS）的二次电子成像（SEI）模式下，在扫描电子显微镜下定位和识别与细胞表面结合的金颗粒。这使得能够可靠地识别金颗粒，同时获得高质量的细胞显微照片。以伴刀豆球蛋白A（ConA）和麦胚凝集素（WGA）这两种凝集素的受体在培养的Raji细胞和人红细胞表面的分布为例进行了说明。Raji细胞和红细胞先用戊二醛固定，再用戊二醛 - 单宁酸混合物后固定，然后与包被有ConA或WGA的金颗粒一起孵育。脱水和临界点干燥后，将样品滤膜安装在铜短柱上并进行碳涂层。在JEOL TEMSCAN 100CX II电子显微镜上检查细胞。金颗粒可以用EDS分析仪识别，当电子束以100,000倍或更高的放大倍数聚焦在单个金颗粒上时，该分析仪能够检测到金的光谱。在高达100,000倍的相同放大倍数下获得了相同细胞的高分辨率照片。
