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J Mol Diagn. 2019 Jul;21(4):658-676. doi: 10.1016/j.jmoldx.2019.03.006. Epub 2019 May 2.

Multilaboratory Assessment of a New Reference Material for Quality Assurance of Cell-Free Tumor DNA Measurements.

Author information

1
Material Measurement Laboratory, National Institute of Standards and Technology, Gaithersburg, Maryland.
2
SeraCare Life Sciences, Inc., Milford, Massachusetts.
3
Frederick National Laboratory for Cancer Research, National Cancer Institute, Leidos Biomedical Research, Inc., Frederick, Maryland.
4
Boston University School of Medicine, Boston, Massachusetts.
5
Johns Hopkins University School of Medicine, Baltimore, Maryland.
6
University of North Carolina School of Medicine, Chapel Hill, North Carolina.
7
Asuragen, Inc., Austin, Texas.
8
Archer Diagnostics, Inc., Boulder, Colorado.
9
Department of Pathology and Genetics, Sahlgrenska Cancer Center, Institute of Biomedicine, Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden; Wallenberg Centre for Molecular and Translational Medicine, University of Gothenburg, Sweden.
10
Department of Pathology and Genetics, Sahlgrenska Cancer Center, Institute of Biomedicine, Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden.
11
Early Detection Research Network, National Cancer Institute, Bethesda, Maryland.
12
Material Measurement Laboratory, National Institute of Standards and Technology, Gaithersburg, Maryland. Electronic address: kenneth.cole@nist.gov.

Abstract

We conducted a multilaboratory assessment to determine the suitability of a new commercially available reference material with 40 cancer variants in a background of wild-type DNA at four different variant allele frequencies (VAFs): 2%, 0.50%, 0.125%, and 0%. The variants include single nucleotides, insertions, deletions, and two structural variations selected for their clinical importance and to challenge the performance of next-generation sequencing (NGS) methods. Fragmented DNA was formulated to simulate the size distribution of circulating wild-type and tumor DNA in a synthetic plasma matrix. DNA was extracted from these samples and characterized with different methods and multiple laboratories. The various extraction methods had differences in yield, perhaps because of differences in chemistry. Digital PCR assays were used to measure VAFs to compare results from different NGS methods. Comparable VAFs were observed across the different NGS methods. This multilaboratory assessment demonstrates that the new reference material is an appropriate tool to determine the analytical parameters of different measurement methods and to ensure their quality assurance.

PMID:
31055023
PMCID:
PMC6626992
[Available on 2020-07-01]
DOI:
10.1016/j.jmoldx.2019.03.006
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