title: Methacarn fixation--effects of tissue processing and storage conditions on detection of mRNAs and proteins in paraffin-embedded tissues.
abstract: In this study, we examined suitable conditions for tissue fixation with methacarn and ethanol dehydration and storage of paraffin-embedded tissues (PETs) on gene expression analysis. With fixation and dehydration of rat liver tissues for up to 16 h (overnight) and 1 week, respectively, at 4 degrees C, integrity of extracted total RNAs and polypeptides did not vary, the former integrity being constantly lower than that with unfixed frozen tissue, while protein yield was slightly reduced with increasing dehydration. Retained expression levels of mRNAs and proteins were mostly unaffected by the period of fixation but slightly fluctuated with the length of dehydration. When PETs were stored for up to 12 months, integrity of both total RNAs and polypeptides was retained at 4 degrees C but reduced at room temperature. Reduced expression levels of mRNAs and proteins were also noted by storage at room temperature after 12 and 3 months, respectively. However, neither tissue processing nor storage affected variability in either mRNA or protein levels among samples. Thus, the results suggest that, for gene expression analysis, tissues can be fixed with methacarn and dehydrated for at least 1 day and 1 week, respectively, and PETs can be stored for at least 12 months, but a temperature of 4 degrees C is preferable.
authors.affiliation: Division of Pathology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
authors.forename: Kyoung-Youl
authors.initials: KY
authors.lastname: Lee
authors.forename: Makoto
authors.initials: M
authors.lastname: Shibutani
authors.forename: Kaoru
authors.initials: K
authors.lastname: Inoue
authors.forename: Keiko
authors.initials: K
authors.lastname: Kuroiwa
authors.forename: Mami
authors.initials: M
authors.lastname: U
authors.forename: Gye-Hyeong
authors.initials: GH
authors.lastname: Woo
authors.forename: Masao
authors.initials: M
authors.lastname: Hirose
mesh_terms: D019342:Acetic Acid; D000818:Animals; D002725:Chloroform; D008297:Male; D000432:Methanol; D016612:Paraffin Embedding; D011506:Proteins / Q000032:analysis*; D012333:RNA, Messenger / Q000032:analysis*; D051381:Rats
chemical_list: D011506:Proteins; D012333:RNA, Messenger; C046544:methacarn; D002725:Chloroform; D019342:Acetic Acid; D000432:Methanol