Effect of formalin, acetone, and RNAlater fixatives on tissue preservation and different size amplicons by real-time PCR from paraffin-embedded tissue

Csilla Páska, Krisztina Bögi, László Szilák, Annamária Tokés, Erzsébet Szabó, István Sziller, János Rigó, Gábor Sobel, István Szabó, Pál Kaposi-Novák, András Kiss, Zsuzsa Schaff

Research output: Contribution to journalArticle

55 Citations (Scopus)

Abstract

RNA recovered from paraffin-embedded tissue has been reported to be a suitable substrate for polymerase chain reaction. During tissue fixation and paraffin embedding, RNA undergoes degradation, but with certain restrictions, it can be used for gene expression studies. At the same time, formalin-fixed, paraffin embedded histopathology archives contain an unestimable collection, which could be analyzed to investigate changes in mRNA expression in pathologic processes. To decide for future tissue conservation of pathology samples, it would be reasonable to satisfy both histologic and molecular biologic needs. The effect of three different fixation methods, RNAlater (SIGMA R 0901, St Louis, MO), acetone, and formalin, were compared by histology, immunohistochemistry, and real-time PCR. To assess tissue structure preservation and antigenicity, hematoxylin-eosin staining and immunohistochemistry were performed; to assess RNA quality, RNA was extracted and the transcription of different amplicon sizes (121, 225, 406 bp for GAPDH; 166, 310, 536 bp for β globin) were examined on human endometrium samples. The most adequate tissue preservation was found in case of formalin fixation, while there were no significant differences in the three fixatives' yields for various size real-time PCR amplicons. Longer amplicons (above ∼225 bp) have limited use for gene expression studies, while shorter amplicons could give more reliable results.

Original languageEnglish
Pages (from-to)234-240
Number of pages7
JournalDiagnostic Molecular Pathology
Volume13
Issue number4
DOIs
Publication statusPublished - Dec 1 2004

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Keywords

  • Endometrium
  • Formalin fixation
  • RNAlater
  • Real-time PCR

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Molecular Biology
  • Cell Biology

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