GOLD CLUSTER LABELS AND RELATED TECHNOLOGIES IN MOLECULAR MORPHOLOGY.
Although intensely colored, even the largest colloidal gold particles are not, on their own, sufficiently colored for routine use as a light microscopy stain: only with very abundant antigens or with specialized illumination methods can bound gold be seen. Colloidal gold probes were developed primarily as markers for electron microscopy, for which their very high electron density and selectivity for narrow size distributions when prepared in different ways rendered them highly suited. The widespread use of gold labeling for light microscopy was made possible by the introduction of autometallographic enhancement methods. In these processes, the bound gold particles are exposed to a solution containing metal ions and a reducing agent; they catalyze the reduction of the ions, resulting in the deposition of additional metal selectively onto the particles. On the molecular level, the gold particles are enlarged up to 30-100 nm in diameter; on the macroscale level, this results in the formation of a dark stain in regions containing bound gold particles, greatly increasing visibility and contrast. The applications of colloidal gold have been described elsewhere in this chapter, we will focus on the use of covalently linked cluster complexes of gold and other metals. A gold cluster complex is a discrete molecular coordination compound comprising a central core, or ''cluster'' of electron-dense metal atoms, ligated by a shell of small organic molecules (ligands), which are linked to the metal atoms on the surface of the core. This structure gives clusters several important advantages as labels. The capping of the metal surface by ligands prevents non-specific binding to cell and tissue components, which can occur with colloidal gold. Cluster compounds are more stable and may be used under a wider range of conditions. Unlike colloidal gold, clusters do not require additional macromolecules such as bovine serum albumin or polyethylene glycol for stabilization, and the total size of the label is therefore significantly smaller. Since the clusters considered in this chapter are generally less than 3 nm in diameter, this allows the preparation of probes that are much smaller than conventional immunocolloids, and cluster labeling can take advantage of the higher resolution and penetration available with smaller conjugates. Most importantly, while colloidal gold is adsorbed to its conjugate probe, clusters are conjugated by chemically specific covalent cross-linking. Therefore, the range of possible conjugate targeting agents includes any probe containing an appropriate reactive group. Clusters conjugates have been prepared with a wide variety of molecules that do not form colloidal gold conjugates, including lipids, oligonucleotides, peptides, and other small molecules. In addition to the development of gold cluster labeling technology, this chapter will also review new developments in the related metallographic, or metal deposition, methods. This includes gold enhancement, in which gold rather than silver is selectively deposited onto gold particles. We will also describe some results obtained using another novel metallographic procedure, enzyme metallography, in which metal is directly deposited from solution by an enzymatic reaction. Because the original, and most widespread, use of metal cluster labels is in electron microscopy, many of the light microscopy methods described were developed as extensions of, or complements to electron microscopy methods, and demonstrate their greatest advantages when used with electron microscopy; therefore reference will also be made to the electron microscope methods used in the same studies, and the unique information that may be obtained from the correlation of both methods.
- Research Organization:
- BROOKHAVEN NATIONAL LABORATORY (US)
- Sponsoring Organization:
- DOE/WFO:NIH (US)
- DOE Contract Number:
- AC02-98CH10886
- OSTI ID:
- 15008839
- Report Number(s):
- BNL-72415-2004-BC; R&D Project: EB02181(NIH) RR017545 (NIH); 40041200 40041200; TRN: US200427%%405
- Resource Relation:
- Other Information: PBD: 4 Feb 2004; Related Information: ADVANCES IN PTHOLOGY, MICROSCOPY, AND MOLECULAR MORPHOLOGY SERIES: MOLECULAR MORPHOLOGY IN HUMAN TISSUES: TECHNIQUES AND APPLICATIONS, VOL. 2 CHAPTER 4
- Country of Publication:
- United States
- Language:
- English
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