Evaluation of non-formalin tissue fixation for molecular profiling studies

John W. Gillespie*, Carolyn J.M. Best, Verena E. Bichsel, Kristina A. Cole, Susan F. Greenhut, Stephen M. Hewitt, Mamoun Ahram, Yvonne B. Gathright, Maria J. Merino, Robert L. Strausberg, Jonathan I. Epstein, Stanley R. Hamilton, Gallya Gannot, Galina V. Baibakova, Valerie S. Calvert, Michael J. Flaig, Rodrigo F. Chuaqui, Judi C. Herring, John Pfeifer, Emmanuel F. PetricoinW. Marston Linehan, Paul H. Duray, G. Steven Bova, Michael R. Emmert-Buck

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

231 Scopus citations

Abstract

Using a general strategy for evaluating clinical tissue specimens, we found that 70% ethanol fixation and paraffin embedding is a useful method for molecular profiling studies. Human prostate and kidney were used as test tissues. The protein content of the samples was analyzed by one-dimensional gel electrophoresis, immunoblot, two-dimensional gel electrophoresis, and layered expression scanning. In each case, the fixed and embedded tissues produced results similar to that obtained from snap-frozen specimens, although the protein quantity was somewhat decreased. Recovery of mRNA was reduced in both quantity and quality in the ethanol-fixed samples, but was superior to that obtained from formalin-fixed samples and sufficient to perform reverse transcription polymerase chain reactions. Recovery of DNA from ethanol-fixed specimens was superior to formalin-fixed samples as determined by one-dimensional gel electrophoresis and polymerase chain reaction. In conclusion, specimens fixed in 70% ethanol and embedded in paraffin produce good histology and permit recovery of DNA, mRNA, and proteins sufficient for several downstream molecular analyses. Complete protocols and additional discussion of relevant issues are available on an accompanying website (http://cgap-mf.inf.nih.gov/).

Original languageEnglish
Pages (from-to)449-457
Number of pages9
JournalAmerican Journal of Pathology
Volume160
Issue number2
DOIs
StatePublished - 2002
Externally publishedYes

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