Treatment of tissue sections for in situ hybridization.
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.
Tesch
Gregory H
GH
Department of Nephrology, Monash University, Monash Medial Center, Melbourne, Australia.
Lan
Hui Y
HY
Nikolic-Paterson
David J
DJ
D000818:Animals; D019286:Endopeptidase K / Q000378:metabolism; D005612:Freeze Drying; D006652:Histological Techniques; D007150:Immunohistochemistry; D017403:In Situ Hybridization / Q000379:methods*; D008867:Microtomy; D008872:Microwaves; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis*; D016610:Tissue Embedding; D016707:Tissue Fixation
D012333:RNA, Messenger; D019286:Endopeptidase K