Multicentre validation study of nucleic acids extraction from FFPE tissues.

Details

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State: Public
Version: Final published version
Serval ID
serval:BIB_861ACDAE7203
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Multicentre validation study of nucleic acids extraction from FFPE tissues.
Journal
Virchows Archiv : an international journal of pathology
Author(s)
Bonin S., Hlubek F., Benhattar J., Denkert C., Dietel M., Fernandez P.L., Höfler G., Kothmaier H., Kruslin B., Mazzanti C.M., Perren A., Popper H., Scarpa A., Soares P., Stanta G., Groenen P.J.
ISSN
1432-2307 (Electronic)
ISSN-L
0945-6317
Publication state
Published
Issued date
09/2010
Peer-reviewed
Oui
Volume
457
Number
3
Pages
309-317
Language
english
Notes
Publication types: Journal Article ; Multicenter Study ; Research Support, Non-U.S. Gov't ; Validation Studies
Publication Status: ppublish
Abstract
In most pathology laboratories worldwide, formalin-fixed paraffin embedded (FFPE) samples are the only tissue specimens available for routine diagnostics. Although commercial kits for diagnostic molecular pathology testing are becoming available, most of the current diagnostic tests are laboratory-based assays. Thus, there is a need for standardized procedures in molecular pathology, starting from the extraction of nucleic acids. To evaluate the current methods for extracting nucleic acids from FFPE tissues, 13 European laboratories, participating to the European FP6 program IMPACTS (www.impactsnetwork.eu), isolated nucleic acids from four diagnostic FFPE tissues using their routine methods, followed by quality assessment. The DNA-extraction protocols ranged from homemade protocols to commercial kits. Except for one homemade protocol, the majority gave comparable results in terms of the quality of the extracted DNA measured by the ability to amplify differently sized control gene fragments by PCR. For array-applications or tests that require an accurately determined DNA-input, we recommend using silica based adsorption columns for DNA recovery. For RNA extractions, the best results were obtained using chromatography column based commercial kits, which resulted in the highest quantity and best assayable RNA. Quality testing using RT-PCR gave successful amplification of 200 bp-250 bp PCR products from most tested tissues. Modifications of the proteinase-K digestion time led to better results, even when commercial kits were applied. The results of the study emphasize the need for quality control of the nucleic acid extracts with standardised methods to prevent false negative results and to allow data comparison among different diagnostic laboratories.

Keywords
Formaldehyde, Humans, Nucleic Acids/analysis, Nucleic Acids/isolation & purification, Paraffin Embedding, Pathology, Molecular/methods, Pathology, Molecular/standards, Reproducibility of Results, Reverse Transcriptase Polymerase Chain Reaction, Tissue Fixation
Pubmed
Web of science
Open Access
Yes
Create date
07/10/2010 9:34
Last modification date
20/08/2019 14:45
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