Summary
To the casual observer,in situ hybridization (ISH) suggests a method to detect and localize nucleic acid sequences in cell preparations, tissue sections or chromosome spreads. This paper is an attempt to demonstrate that ISH is very much more versatile than is immediately apparent; a number of possible alternatives in different steps in the procedure have been defined and specific methods recommended that are known to be successful. The text is limited to the detection of viral nucleic acids in tissue sections and cells, although many of the concepts may be applied more generally. Methods and practical details have been reduced to a minimum as they are covered in the accompanying paper by Dr. F. Negro et al. (see page 177 of this issue) and in references. A comprehensive review of ISH procedures has been compiled recently12.
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References
Arrand J. E.: Preparation of nucleic acid probes. In:Hames B.D., Higgins S.J. (Eds): Nucleic acid hybridization. IRL Press, Oxford, 1986; p. 17.
Betrm H. E., Haase A. T., Vyas G. N.: Molecular pathogenesis of hepatitis B virus infection: simultaneous detection of viral DNA and antigens in paraffin-embedded liver sections - Lancetii, 771, 1984.
Britten R.J., Davidson E. H.: Hybridization strategy. In:Hames B.D., Higgins S.J. (Eds): Nucleic acid hybridization. IRL Press, Oxford, 1986; p. 3.
Cox K., Deleon D.V., Angerer L. M., Angerer R. C.: Detection of mRNAs in sea urchin embryos byin situ hybridization using asymmetric RNA probes - Develop. Biol.101, 485, 1984.
Feinberg A.P., Vogelstein B.: A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity - Analyt. Biochem.132, 6, 1983.
Gendelman H.E., Moench T. R., Narayen O., Griffin D. E., Clements J. R.: A double labelling technique for performing immunochemistry andin situ hybridization in virus-infected cell cultures and tissues-J. virol. Meth.11, 93, 1985.
Gowans E.J.,Baroudy B.M.,Negro E.,Ponzetto A.,Purcell R.H.,Gerin J.L.: Evidence for replication of hepatitis delta virus RNA in hepatocyte nuclei after in vivo infection - Virology165, 1988. (In press).
Gowans E.J., Borrell C.J.: Widespread presence of cytoplasmic HBcAg in hepatitis B infected liver detected by improved immunochemical methods -J. clin. Pathol.38, 393, 1985.
Gowans E.J., Burrell C.J., Jilbert A.R., Marmion B.P.: Detection of hepatitis B virus DNA sequences in infected hepatocytes byin situ cytohybridization -J. reed. Virol.8, 67, 1981.
Gowans E.J., Burrell C.J., Jilbert A.R., Marmion B.P.: Patterns of single- and doublestranded hepatitis B virus DNA and viral antigen accumulation in infected liver cells -J. gen. Virol.64, 1229, 1983.
Gowans E.J., Borrell C.J., Jilbert A.R., Marmion B.E.: Cytoplasmic (but not nuclear) hepatitis B virus (HBV) core antigen reflects HBV-DNA synthesis at the level of the infected hepatocyte - Intervirology24, 220, 1985.
Gowans E.J., Jilbert A.R., Burrell C.J.: Detection of specific DNA and RNA sequences in tissues and cells byin situ hybridization. In:Symons R.H. (Ed.): Nucleic acid probes. CRC Press Inc., Boca Raton, 1988. (In press).
Gowans E.J., Negro E, Baroudy B.M., Korba B.E., Bonino F., Ponzetto A., Purcell R.H., Gerin J.L.: Molecular markers of WHV and HDV in single cells. In:Robinson W., Koike K., Will H. (Eds): Hepadnaviruses. Alan R. Liss, New York, 1987; p. 591.
Haase A., Brahic M., Stowring L., Blum H.: Detection of viral nucleic acids byin situ hybridization - Meth. Virol.7, 189, 1984.
Jilbert A.R., Burrell C.J., Gowans E.J., Rowland A.R.: Histological aspects ofin situ hybridization: detection of poly(A) nucleotide sequences in mouse liver sections as a model system - Histochemistry85, 505, 1986.
Jilbert A.R., Freiman J.S., Gowans E.J., Holmes M., Cossart Y.E., Burrell C.J.: Duck hepatitis B virus DNA in liver, spleen and pancreas: analysis byin situ and Southern blot hybridization -Virology158, 330, 1987.
Korba B.E.,Gowans E.J.,Welts E.V.,Tennant B.C.,Clarke R.,Gerin J.L.: Systemic distribution of woodchuck hepatitis virus in the tissues of experimentally-infected animals - Virology165, 1988. (In press).
Lawrence J.B., Singer R.H.: Intracellular localization of messenger RNAs for cytoskeletal proteins - Cell45, 407, 1986.
Maddox P.H., Jenkins D.: 3-aminopropyltriethoxysilane (APES): a new advance in section adhesion -J. clin. Pathol.40, 1256, 1987.
MCallister L.B., Scheller R.H., Kandel E.R., Axel R.: Insitu hybridization to study the origin and fate of identified neurons - Science222, 800, 1983.
Melton D.A., Krieg P.A., Rebagliati M.R., Maniatis W., Zinn K., Green M.R.: Efficientin vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter - Nucleic Acids Res.12, 7035, 1984.
Pardue M.L.:In situ hybridization. In:Hames B. D., Higgins S.J. (Eds): Nucleic acid hybridization. IRL Press, Oxford, 1986; p. 179.
Renz M., Kurz C.: A colorimetric method for DNA hybridization - Nucleic Acids Res.12, 3435, 1984.
Rogers A.W.: Techniques of autoradiography. Elsevier/North-Holland, Amsterdam, 1981; 3rd ed.
Taylor J., Mason W., Summers J., Goldberg J., Aldrich C., COATES L., GERIN J., GOWANS E.: Replication of human hepatitis delta virus in primary cultures of woodchuck hepatocytes - J. Virol.61, 2891, 1987.
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Gowans, E.J. Strategies forin situ hybridization. La Ricerca Clin. Lab. 18, 171–176 (1988). https://doi.org/10.1007/BF02918882
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DOI: https://doi.org/10.1007/BF02918882