Functional DNA quantification guides accurate next-generation sequencing mutation detection in formalin-fixed, paraffin-embedded tumor biopsies

The formalin-fixed, paraffin-embedded (FFPE) biopsy is a challenging sample for molecular assays such as targeted next-generation sequencing (NGS). We compared three methods for FFPE DNA quantification, including a novel PCR assay (‘QFI-PCR’) that measures the absolute copy number of amplifiable DNA...

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Published inGenome medicine Vol. 5; no. 8; p. 77
Main Authors Sah, Sachin, Chen, Liangjing, Houghton, Jeffrey, Kemppainen, Jon, Marko, Adam C, Zeigler, Robert, Latham, Gary J
Format Journal Article
LanguageEnglish
Published London BioMed Central 30.08.2013
BioMed Central Ltd
Subjects
4. Proteomics & metabolomics in medicine
Bioinformatics
Biomedical and Life Sciences
Biomedicine
Cancer Research
DNA
Formaldehyde
Human Genetics
Medicine/Public Health
Metabolomics
Method
Systems Biology
Tumors
FFPE Sample
Qubit Assay
Quality Control Metrics
PIK3CA H1047R
Haploid Copy
Online AccessGet full text
ISSN1756-994X
1756-994X
DOI10.1186/gm481

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Summary:The formalin-fixed, paraffin-embedded (FFPE) biopsy is a challenging sample for molecular assays such as targeted next-generation sequencing (NGS). We compared three methods for FFPE DNA quantification, including a novel PCR assay (‘QFI-PCR’) that measures the absolute copy number of amplifiable DNA, across 165 residual clinical specimens. The results reveal the limitations of commonly used approaches, and demonstrate the value of an integrated workflow using QFI-PCR to improve the accuracy of NGS mutation detection and guide changes in input that can rescue low quality FFPE DNA. These findings address a growing need for improved quality measures in NGS-based patient testing.
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ISSN:1756-994X
1756-994X
DOI:10.1186/gm481