Summary
The treatment of tissue sections to enhance probe access to target mRNA is a critical step in the methodology of in situ hybridization. We have overcome some of the problems encountered in enzyme-based treatment of tissue sections by the application of microwave oven heating. Microwave treatment can (1) replace proteinase K digestion for frozen sections; (2) enhance proteinase K digestion in paraffin sections; (3) denature mRNA structure to enable better probe access; (4) preserve tissue architecture; and (5) inactivate endogenous alkaline phosphatase within tissue sections to reduce background with immunohistochemistry-based probe detection.
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References
Lan H. Y., Mu W., Ng Y. Y., Nikolic-Paterson D. J., and Atkins R. C. (1996) A simple, reliable and sensitive method for nonradioactive in situ hybridization: use of microwave heating to improve hybridization efficiency and preserve tissue morphology. J. Histochem. Cytochem. 44, 281–287.
Lan H. Y., Mu W., Nikolic-Paterson D. J., and Atkins R. C. (1996) The applications of microwave techniques in multiple immunostaining and in situ hybridization. Nephrology 2, S116–S121.
Lan H. Y., Mu W., Nikolic-Paterson D. J., and Atkins R. C. (1995) A novel, simple, reliable and sensitive method of multiple immunoenzymic staining: use of microwave oven heating to block antibody cross-reactivity and retrieve antigens. J. Histochem. Cytochem. 43, 97–102.
Gu J., Farley R., Shi S-R., and Taylor C. R. (2000) Target retrieval for in situ hybridization, in Antigen Retrieval Techniques: Immunohistochemistry and Molecular Morphology (Shi S-R, Gu J., Taylor C. R. eds.), Eaton Publishing, Natick, MA, pp. 115–128.
Lan H. Y., Yu X. Q., Yang N., et al. (1998) De novo glomerular osteopontin expression in rat crescentic glomerulonephritis. Kidney Int. 53, 136–145.
Tesch G. H., Nikolic-Paterson D. J., Metz C. N., et al. (1998) Rat mesangial cells express macrophage migration inhibitory factor in vitro and in vivo. J. Am. Soc. Nephrol. 9, 417–424.
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Tesch, G.H., Lan, H.Y., Nikolic-Paterson, D.J. (2006). Treatment of Tissue Sections for In Situ Hybridization. In: Darby, I.A., Hewitson, T.D. (eds) In Situ Hybridization Protocols. Methods in Molecular Biology™, vol 326. Humana Press. https://doi.org/10.1385/1-59745-007-3:1
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DOI: https://doi.org/10.1385/1-59745-007-3:1
Publisher Name: Humana Press
Print ISBN: 978-1-58829-402-9
Online ISBN: 978-1-59745-007-2
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