A simple and rapid technique to process formalin-fixed, paraffin-embedded tissues for the detection of viruses by the polymerase chain reaction.

Author(s): Kiene P, Milde-Langosch K, Runkel M, Schulz K, Löning T

Publication: Virchows Arch A Pathol Anat Histopathol, 1992, Vol. 420, Page 269-73

PubMed ID: 1313196 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 deparaffinization, proteinase k digestion, and DNA extraction procedure on the amplification of viral DNA from FFPE heart, lung, kidney, placental, pharynx, and vaginal autopsy and biopsy specimens. The effect of specimen age and the method used to detect the PCR products were also examined.

   Summary of Findings:

   Amplification of products up to 450 bp long was successful when the specimens were boiled for 3-5 minutes in chelating resin sodium form or digested in proteinase K and extracted by phenol-chloroform. In contrast, no amplification was observed in specimens that were deparaffinized in xylene with or without subsequent proteinase k digestion or in specimens boiled in water. The authors report that amplification was successful regardless of storage duration and that there was no effect of hybridization probe type.

   Biospecimens

   • Tissue - Heart
   • Tissue - Lung
   • Tissue - Vagina
   • Tissue - Placenta
   • Tissue - Nasopharynx
   • Tissue - Kidney

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma
   • AIDS/HIV-related
   • Cytomegalovirus
   • Autopsy
   • Other diagnoses

   Platform:

   Analyte	Technology Platform
   DNA	PCR
   DNA	Southern blot

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Protein digestion	None Proteinase K at 55 degrees C for 3 h
   Analyte Extraction and Purification	Deparaffinization	Boiling in chelating resin sodium form followed by centrifugation None Xylene Boiling in water
   Analyte Extraction and Purification	Analyte isolation method	None Phenol-chloroform extraction
   Storage	Storage duration	Short 12 years
   PCR Specific	Targeted nucleic acid	E6 region of human papilomavirus Human papilomavirus open reading frame region L1 IE region of cytomegalovirus Epstein Bar virus Beta- globin
   PCR Specific	Length of gene fragment	120 bp 137 bp 450 bp
   Southern blot Specific	Detection method	P32 labeled probe Digoxigenin labeled probe
   Analyte Extraction and Purification	Rehydration of dried sample/specimen	1% N-lauroylsarcosine in TE Water

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