Electron microscopy in situ hybridization

Chevalier J, Yi J, Michel O, Tang X-M. 1997. Biotin and digoxigenin as labels for light and electron microscopy in situ hybridization process where do we stand J Histochem Cytochem 45 481-491. 

McFadden G. In situ hybridization techniques molecular cytology goes ultrastruc-tural, in Electron Microscopy of Plant Cells (Hall J, Hawes C, eds.), Academic Press, London, 1991, pp. 219-255. 

Electron microscopy permits the detailed study of cell relationships within tissues and organelles within cells. Two ultrastructural techniques in which gold probes have proven invaluable are mununocytochemistry and in situ hybridization, and these will be described here. Until 1980, peroxidase was the marker of choice, but now colloidal gold is almost universally used. The advantages of colloidal gold are that ... 

Staining of consecutive sections with histochemical reagents for protein and antibodies against myrosinase showed that myrosin cells actually contain myrosinase (Fig. 4.2D,E).18,43,44 By electron microscopy immunogold labelling studies, the enzyme was localized to the interior of the vacuoles of the myrosin cells (Fig. 4.2F).18,40,41 It has also been shown by several in situ hybridization experiments that myrosinase transcripts are located in these cells (Fig. 4.2G),6 demonstrating that the cellular localization of myrosinase is due to transcriptional regulation and not to a transport process. No evidence for transport of myrosinase currently exists. These methods for detection and identification of myrosin cells and myrosinase expression are illustrated (Fig. 4.2). For a historical overview of myrosinase localization studies, see Bones and Rossiter.45... 

Chronwall BM, Hook GR, Millington WR (1988) Dopaminergic regulation of the biosynthetic activity of individual melanotropes in the rat pituitary intermediate lobe a morphometric analysis by light and electron microscopy and in situ hybridization. Endocrinology 725 1992-2002. 

Cowdry type A bodies of HSV are not easy to demonstrate in small brain-biopsy specimens. This argues for sensitive and specific methods of identification such as in situ hybridization (ISH), PCR, and IHC. s Electron microscopy may demonstrate viral particles within the nuclei or cytoplasm but is less sensitive and less specific. Culture and sequential serologic CSF evaluations are slow but are still the most accurate methods of diagnosis for many viral CNS infections, including HSV. 

N. J. Hutchison, Hybridisation Histochemistry In Situ Hybridization at the Electron Microscope Level, in Immunolabeling for Electron Microscopy (eds. J. M. Polak and I. M. Varndell), Elsevier, Amsterdam, 1984, pp. 341-351. 

Spiess, E., Zimmermann, H.-P., and LUnsdorf, H. (1987). Negative staining of protein molecules and filaments. In Electron Microscopy in Molecular Biology A Practical Approach (J. Sommer-ville and U. Scheer, eds.), pp. 147-166. IRL Press, Oxford. UK and Washington, DC. Spyropoulos, B., and Moens, P. B. (1994). In situ hybridization of meiotic prophase chromosomes. Methods Mol. Biol. 33, 131-139. 

Deerinck, T. J., Martone, M. E., Lev-Ram, V., Green, D. P., Tsien, R. Y., Spector, D. L., Huang, S., and Ellisman, M. H. (1994). Fluorescence photooxidation with eosin A method for high resolution immunolocalization and in situ hybridization detection for light and electron microscopy. J. Cell Biol. 126, 901-910. 

Methods BF = bright field LM of tissue impression smears, wet-mounts, stained whole mounts LM = light microscopy PH = phase microscopy DF = dark-field microscopy TEM/SEM = transmission/scanning electron microscopy of sections or of purified or semi-purified virus ELISA = enzyme-linked immunosorbent assay PAb = polyclonal antibodies MAb = monoclonal antibodies DBH = dot blot hybridization ISH = in situ hybridization PCR = polymerase chain reaction RT-PCR = reverse transcription PCR. 

Pig. 10.13 Future directions of hot electron studies, including (a) development of hybrid nanoparticle-nanodiode systems, and (b) in situ surface characterization. The cartoon depicts the conductive atomic force microscopy experiments on Au/liOj nanostructures under exothermic catalytic reactions or photon irradiation... 

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