Colloidal gold as a cytochemical marker in electron microscopy

Abstract

Colloidal gold is orange to red, displays electron-opaque properties and is capable of strong emission of secondary electrons. As gold particles can be produced in different sizes and can be labelled with macromolecules which keep their specific properties, gold markers have found ases in light and fluorescent microscopy, and many applications in scanning and transmission electron microscopy.In biology, the most sophisticated analysis performcd in vitro will never suppress the urge of an investigator to visualize the exact location of a cell component.From the time the Dutch naturalist Antonie van Leeuwenhoek (1632-1723) made his first discoveries with a simple instrument, microscopy has indeed proved to be a powerful tooi for understanding the microcosm. Thanks to patient efforts and to an amazing skill in using lenses, van Leeuwenhoek was able to obtain results which, at the time, were thought to be truly marvellous. His experiments became so popular that it was fashionable in the upper class society of Delft to Eind distraction, satisfaction and even joy in observing nature through his primitive microscopes. These inner feelings are still strongly feit by modern researchers using much more powerful instruments.Clearly, van Leeuwenhoek was a self-taught microscopist, but his observations led to many discoveries of great importante. He was the first to observe protozoa, spermatozoids and red blood cells, and the first representation of a bacterium is to be found in one of his drawings of 1682.The development of optical microscopes has continued since that time, but the limits imposed by the wavelengths of visible light prompted the search for high resolution techniques. The first transmission electron microscopes having better resolution than light microscopes were built in 1932. Thirty years later, the scanning electron microscope was introduccd. Scanning electron microscopy (SEM) gives a three-dimensional quality to specimen images. Normally, the instrument is operated by scanning, or sweeping, a very narrow beam of electrons back and forth across a metal-coated specimen, revealing its surface features rather than its internal structure.The possibility of localizing specific tissue components attracted the early microscopists, but it was Raspail (1794-1878), a French botanist, pharmacist, microscopist and politician, who first used the microscope to study the chemical reactions of tissue materials. He is considered to be the founder of histochemistry and some of the reactions which he discovered are still applied today. Over the years, a large number of techniques has been developed to identify, locate and quantify specific tissues and cell components. Among the methods having a narrow specificity, the use of fluorescent antibodies was introduced in 1941 by Coons et al. (1). A great advance was then achieved at the ultrastructural level with the development of enzyme cytochemistry (2, 3, 4) and the application of particulate markers, such as ferritin, conjugated to antibodies (5, 6). Ferritin is a large molec...

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

“…The resolution obtainable by such particulate tracers is superior to cytochemical methods involving enzymes such as peroxidase (Courtoy et al, 1983). Furthermore, gold particles of various sizes can be easily produced (Horisberger, 1979;Geuze, 1981 , 1985;Roth, 1982b) and, owing to the great size homogeneity of the particle populations, double-labelling experiments can be performed on the same preparation.…”

Photosystem II and cytochromeb-559 in the stroma lamellae of barley chloroplasts

“…For immunocytochemistry on thin sections, chloroplasts were fixed, embedded in Lowicryl K4M and labelled as previously described (49). Gold sols (12 nm) were prepared according to (22) and conjugated with protein A for detection of pAb's or with a rabbit anti-mouse IgG antibody (DAKO Immunoglobulins A/S, Copenhagen) for detection of mAb's. Quantitation of labelling was done by counting gold particles over appressed thylakoid regions versus non-appressed regions (stroma lamellae and grana end lamellae).…”

Use of colloidal gold particles in double-labeling immunoelectron microscopy of ultrathin frozen tissue sections.

“…Secondly, the coating of the gold particles with PA is rapid and uncomplicated, yielding a probe that is suitable to label IgG of most mammals, except goat and sheep (19). Thirdly, various sizes of gold particles can be made for highresolution studies (13).' This opens up the field of multiple labeling of antigens.…”