Multicentre validation study of nucleic acids extraction from FFPE tissues.

Author(s): Bonin S, Hlubek F, Benhattar J, Denkert C, Dietel M, Fernandez PL, Höfler G, Kothmaier H, Kruslin B, Mazzanti CM, Perren A, Popper H, Scarpa A, Soares P, Stanta G, Groenen PJ

Publication: Virchows Arch, 2010, Vol. 457, Page 309-17

PubMed ID: 20665046 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 RNA and DNA extraction methods on the amplification of colorectal, lung, and ovary FFPE tumor specimens.

   Summary of Findings:

   The highest DNA yield was obtained when no further purification steps were taken after the initial DNA extraction. There was no correlation between extraction method and PCR success. The lung tissue allowed for amplification of a 300 bp product, regardless of extraction method, and a 400 bp product was amplifiable in some cases, but the largest amplification product from ovary and colon tissue was 200 bp. The highest quality RNA was from specimens purified using silica based column purification and was especially good using the Qiagen RNeasy FFPE kit, but quality depended on the user. The quality of the RNA did not predict amplification success, regardless of purification method.

   Biospecimens

   • Tissue - Lung
   • Tissue - Colorectal
   • Tissue - Ovary

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   DNA	PCR
   RNA	RT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Analyte isolation method	DNA extraction kit with precipitation step DNA extraction kit without prrecipitation step DNA extraction kit with silica columns RNA extraction with homemade phenol-chloroform protocol RNA extraction with phenol-chloroform based kit RNA extraction with silica based column purification
   Biospecimen Acquisition	Biospecimen location	Lung Ovary Colon
   PCR Specific	Length of gene fragment	100 bp 200 bp 300 bp 400 bp
   RT-PCR Specific	Length of gene fragment	103 bp 150 bp 200 bp 250 bp
   RT-PCR Specific	Targeted nucleic acid	B-actin PBGD B2M
   PCR Specific	Targeted nucleic acid	BIOMED

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