Clinical investigational studies for validation of a next-generation sequencing in vitro diagnostic device for cystic fibrosis testing

Daniel S. Grosu, Lynda Hague, Manjula Chelliserry, Kristina M. Kruglyak, Ross Lenta, Brandy Klotzle, Jonathan San, Wendy M. Goldstein, Sharmili Moturi, Patricia Devers, Julie Woolworth, Eric Peters, Barbara Elashoff, Jay Stoerker, Daynna J. Wolff, Kenneth J. Friedman, W. Edward Highsmith, Erick Lin, Frank S. Ong

Research output: Contribution to journalArticlepeer-review

16 Scopus citations

Abstract

Purpose: Clinical investigational studies were conducted to demonstrate the accuracy and reproducibility of the Illumina MiSeqDx CF System, a next-generation sequencing (NGS) in vitro diagnostic device for cystic fibrosis testing. Methods: Two NGS assays-a Clinical Sequencing Assay (Sequencing Assay) and a 139-Variant Assay (Variant Assay)-were evaluated in both an Accuracy Study and a Reproducibility Study, with comparison to bi-directional Sanger sequencing and PCR as reference methods. For each study, positive agreement (PA), negative agreement (NA), and overall agreement (OA) were evaluated. Results: In the Accuracy Study, the Sequencing Assay achieved PA of 99.7% including the polyTG/polyT region and PA of 100% excluding the region. The Variant Assay achieved PA of 100%. NA and OA were >99.99% for both Assays. In the Reproducibility Study, the Sequencing Assay achieved PA of 99.2%; NA and OA were both 99.7%. The Variant Assay achieved PA of 99.8%; NA and OA were both 99.9%. Sample pass rates were 99.7% in both studies for both assays. Conclusion: This is the first systematic evaluation of a NGS platform for broad clinical use as an in vitro diagnostic, including accuracy validation with multiple reference methods and reproducibility validation at multiple clinical sites. These NGS-based Assays had accurate and reproducible results which were comparable to or better than other methods currently in clinical use for clinical genetic testing of cystic fibrosis.

Original languageEnglish (US)
Pages (from-to)605-622
Number of pages18
JournalExpert Review of Molecular Diagnostics
Volume14
Issue number5
DOIs
StatePublished - Jun 2014

Keywords

  • Clinical validation
  • Cystic fibrosis
  • In vitro diagnostics
  • Next-generation sequencing
  • Sequencing-by-synthesis

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Molecular Medicine
  • Molecular Biology
  • Genetics

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