title: Evaluation of non-formalin tissue fixation for molecular profiling studies.
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.nih.gov/).
authors.affiliation: Pathogenetics Unit, Laboratory of Pathology, National Cancer Institute, Rm. 2C500. Bldg. 10, 9000 Rockville Pike, Bethesda, MD 20892, USA.
authors.forename: John W
authors.initials: JW
authors.lastname: Gillespie
authors.forename: Carolyn J M
authors.initials: CJ
authors.lastname: Best
authors.forename: Verena E
authors.initials: VE
authors.lastname: Bichsel
authors.forename: Kristina A
authors.initials: KA
authors.lastname: Cole
authors.forename: Susan F
authors.initials: SF
authors.lastname: Greenhut
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Hewitt
authors.forename: Mamoun
authors.initials: M
authors.lastname: Ahram
authors.forename: Yvonne B
authors.initials: YB
authors.lastname: Gathright
authors.forename: Maria J
authors.initials: MJ
authors.lastname: Merino
authors.forename: Robert L
authors.initials: RL
authors.lastname: Strausberg
authors.forename: Jonathan I
authors.initials: JI
authors.lastname: Epstein
authors.forename: Stanley R
authors.initials: SR
authors.lastname: Hamilton
authors.forename: Gallya
authors.initials: G
authors.lastname: Gannot
authors.forename: Galina V
authors.initials: GV
authors.lastname: Baibakova
authors.forename: Valerie S
authors.initials: VS
authors.lastname: Calvert
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Flaig
authors.forename: Rodrigo F
authors.initials: RF
authors.lastname: Chuaqui
authors.forename: Judi C
authors.initials: JC
authors.lastname: Herring
authors.forename: John
authors.initials: J
authors.lastname: Pfeifer
authors.forename: Emmanuel F
authors.initials: EF
authors.lastname: Petricoin
authors.forename: W Marston
authors.initials: WM
authors.lastname: Linehan
authors.forename: Paul H
authors.initials: PH
authors.lastname: Duray
authors.forename: G Steven
authors.initials: GS
authors.lastname: Bova
authors.forename: Michael R
authors.initials: MR
authors.lastname: Emmert-Buck
mesh_terms: D000199:Actins / Q000032:analysis / Q000235:genetics; D004247:DNA / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D000431:Ethanol; D005069:Evaluation Studies as Topic; D005404:Fixatives / Q000737:chemistry; D006801:Humans; D007150:Immunohistochemistry; D007668:Kidney / Q000737:chemistry* / Q000166:cytology; D008297:Male; D016612:Paraffin Embedding; D011467:Prostate / Q000737:chemistry* / Q000166:cytology; D017430:Prostate-Specific Antigen / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D012313:RNA / Q000032:analysis*; D016707:Tissue Fixation / Q000379:methods*
chemical_list: D000199:Actins; D005404:Fixatives; D011506:Proteins; D000431:Ethanol; D012313:RNA; D004247:DNA; D017430:Prostate-Specific Antigen