Cell-Free DNA Assay Validations

Publication Date: October 6, 2023
Last Updated: October 12, 2023

Summary of Recommendations for Clinical ctDNA Testing, Reporting, and Publications

  1. Laboratories should clearly define and describe during validation the medical indication for an assay, including the clinical scenarios to which the performance characteristics apply and the methods used.
  2. Laboratories should clearly define and describe during validation clinical assay performance characteristics (sensitivity, specificity, positive predictive value, negative predictive value, accuracy, and concordance) appropriate for the medical indication for the test.
  3. Laboratories should evaluate and describe during validation key clinical assay performance characteristics on an individual variant basis, but these may be aggregated for each variant class. Note: variant classes could include SNVs, indel, copy number alteration, structural variant (eg, fusion), or signature (microsatellite instability, homologous recombination deficiency, or tumor mutational burden).
  4. Laboratories should define and describe during validation the analytical sensitivity (limit of detection) of the assay/s for each variant and/or variant class.
  5. Laboratories should define and describe during validation both in vivo and in vitro potential sources of assay interference.
  6. Laboratories should evaluate and address during validation both in vivo and in vitro potential sources of result interpretation error.
  7. Laboratories should define and describe during validation orthogonal method confirmations, to include the parameters above.
  8. Laboratory reporting for clinical ctDNA assays should describe pre-analytical variables pertinent to both pathologist and clinician understanding of reported results. Note: these may include but are not limited to volume and type of collected fluid, collection tube, and details of storage and processing (eg, refrigeration and centrifugation).
  9. Laboratory reporting for clinical ctDNA assays should describe analytical variables pertinent to both pathologist and provider understanding of reported results. Note: these may include but are not limited to nucleic acid extraction method, library preparation method, volume/concentration of nucleic acid extracted, volume/concentration of nucleic acid used in assay, minimum/mean sequencing depth (for NGS) and droplet scoring parameters (for ddPCR), and details of the bioinformatic pathway.
  10. Laboratory reporting for clinical ctDNA assays should state the analytical sensitivity (limit of detection) of the assay/s for each variant and/or variant class.
  11. Laboratory reporting for clinical ctDNA assays should describe interpretive variables pertinent to both pathologist and provider understanding of reported results. Note: these may include but are not limited to methods employed for variant/signature annotation/interpretation and cutoffs.
  12. Although universal orthogonal confirmation is not required, laboratory reporting for clinical ctDNA assays should have and provide information regarding the laboratory's policy for orthogonal method confirmations and specify any exceptions that may apply.
  13. Publications describing ctDNA assays intended for clinical applications, including description of clinical validation, should include key performance characteristics.
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Overview

Title

Cell-Free DNA Assay Validations

Authoring Organizations