Brief Communication abstract
Nature Methods 6, 139 - 141 (2009)
Published online: 11 January 2009 | doi:10.1038/nmeth.1294
miRNA in situ hybridization in formaldehyde and EDC–fixed tissues
John T G Pena1,2, Cherin Sohn-Lee1, Sara H Rouhanifard1, Janos Ludwig1,4, Markus Hafner1, Aleksandra Mihailovic1, Cindy Lim1, Daniel Holoch1, Philipp Berninger3, Mihaela Zavolan3 & Thomas Tuschl1
MicroRNAs are small regulatory RNAs with many biological functions and disease associations. We showed that in situ hybridization (ISH) using conventional formaldehyde fixation results in substantial microRNA loss from mouse tissue sections, which can be prevented by fixation with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide that irreversibly immobilizes the microRNA at its 5' phosphate. We determined optimal hybridization parameters for 130 locked nucleic acid probes by recording nucleic acid melting temperature during ISH.
- Howard Hughes Medical Institute, Laboratory of RNA Molecular Biology, The Rockefeller University, 1230 York Avenue, Box 186, New York, New York 10065, USA.
- Weill Medical College of Cornell University, 1300 York Avenue, New York, New York 10021, USA.
- Biozentrum, University of Basel and Swiss Institute of Bioinformatics, Klingelbergstrasse 50-70, 4056 Basel, Switzerland.
- Present address: Institute of Clinical Chemistry and Pharmacology, University of Bonn, Sigmund Freud Str. 25, 53127 Bonn, Germany.
Correspondence to: Thomas Tuschl1 e-mail: ttuschl@rockefeller.edu.
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