The effect of tissue decalcification on mRNA retention within bone for in-situ hybridization studies.

Author(s): Walsh L, Freemont AJ, Hoyland JA

Publication: Int J Exp Pathol, 1993, Vol. 74, Page 237-41

PubMed ID: 8392858 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of decalcification agent on morphology and mRNA detection in bone specimens.

   Summary of Findings:

   Decalcification with nitric acid took 2 days, formic acid took 3-4 days, and EDTA took 10 days, as determined by X-ray. Specimens decalcified with EDTA had a similar morphological appearance to specimens that were not decalcified and only a slight reduction in total mRNA signal. In contrast, specimens decalcified in formic or nitric acid showed a loss of morphological details and abnormal H&E staining. Further, in acid-decalcified specimens, the mRNA signal was non-significantly reduced by as much as 50%. Acid-decalcified specimens showed higher background staining, compared to EDTA-decalcified specimens, which was not further reduced by RNAse treatment.

   Biospecimens

   • Tissue - Bone

   Preservative Types

   • Formalin

   Diagnoses:

   • Not specified

   Platform:

   Analyte	Technology Platform
   DNA	In situ hybridization
   Morphology	X-ray
   Morphology	H-and-E microscopy
   Morphology	Light microscopy

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Decalcification solution/ duration	6% nitric acid 90% buffered formic acid 20% EDTA pH 7.4
   In situ hybridization Specific	Targeted nucleic acid	Poly-A tail

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