Non-formalin fixative versus formalin-fixed tissue: A comparison of histology and RNA quality

Daniel Groelz, Leslie Sobin, Philip Branton, Carolyn Compton, Ralf Wyrich, Lynne Rainen

Research output: Contribution to journalArticle

48 Citations (Scopus)

Abstract

Preanalytical handling of tissue samples can influence bioanalyte quality and ultimately outcome of analytical results. The aim of this study was to compare RNA quality, performance in real time RT PCR and histology of formalin-fixed tissue to that of tissue fixed and stabilized with a formalin-free fixative, the PAXgene Tissue System (PAXgene), in an animal model under highly controlled preanalytical conditions. Samples of rat liver, kidney, spleen, intestine, lung, heart muscle, brain, and stomach tissue were either fixed in formalin or fixed in PAXgene or fresh frozen in liquid nitrogen. RNA was extracted from all samples, examined for integrity in microcapillary electrophoresis, and used in a series of quantitative RT PCR assays with increasing amplicon length. Histology of paraffin-embedded samples was determined by staining with hematoxylin and eosin.Histology of all formalin-fixed and PAXgene fixed samples was comparable. RNA with acceptable integrity scores could be isolated from all embedded tissues, 4.0 to 7.2 for formalin and 6.4 to 7.7 for PAXgene, as compared to 8.0 to 9.2 for fresh frozen samples. While RNA with acceptable RINs (RNA integrity number) could be isolated from formalin-fixed samples, in microcapillary electrophoresis this RNA separated with a slower migration rate and displayed diffuse, less focused peaks for ribosomal RNA as compared to RNA from frozen or PAXgene fixed samples. Furthermore, RNA from formalin-fixed tissues exhibited inhibition in quantitative RT PCR assays which increased with increasing amplicon length, while RNA from PAXgene fixed samples did not show such inhibition.In conclusion, our results demonstrate that excluding other preanalytical factors, PAXgene Tissue System preserves histology similarly to formalin, but unlike formalin, does not chemically modify RNA. RNA purified from PAXgene fixed tissues is of high integrity and performs as well as RNA from fresh frozen tissue in RT PCR regardless of amplicon length.

Original languageEnglish (US)
Pages (from-to)188-194
Number of pages7
JournalExperimental and Molecular Pathology
Volume94
Issue number1
DOIs
StatePublished - Feb 2013
Externally publishedYes

Fingerprint

Fixatives
Histology
Formaldehyde
RNA
Tissue
Electrophoresis
Polymerase Chain Reaction
Assays
Ribosomal RNA
Thromboplastin
Liquid nitrogen
Hematoxylin
Eosine Yellowish-(YS)
Liver
Paraffin
Intestines
Muscle
Rats
Real-Time Polymerase Chain Reaction
Brain

Keywords

  • H&E histology
  • PAXgene Tissue
  • Quantitative RT PCR
  • RNA
  • RNA integrity score

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Molecular Biology
  • Pathology and Forensic Medicine

Cite this

Non-formalin fixative versus formalin-fixed tissue : A comparison of histology and RNA quality. / Groelz, Daniel; Sobin, Leslie; Branton, Philip; Compton, Carolyn; Wyrich, Ralf; Rainen, Lynne.

In: Experimental and Molecular Pathology, Vol. 94, No. 1, 02.2013, p. 188-194.

Research output: Contribution to journalArticle

Groelz, Daniel ; Sobin, Leslie ; Branton, Philip ; Compton, Carolyn ; Wyrich, Ralf ; Rainen, Lynne. / Non-formalin fixative versus formalin-fixed tissue : A comparison of histology and RNA quality. In: Experimental and Molecular Pathology. 2013 ; Vol. 94, No. 1. pp. 188-194.
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