Analytical Validation of an Error-Corrected Ultra-Sensitive Ctdna Next-Generation Sequencing Assay | Zendy

Heidi Fettke | Zendy; Jason A. Steen | Zendy; Edmond M. Kwan | Zendy; Patricia Bukczynska | Zendy; Shivakumar Keerthikumar | Zendy; David L. Goode | Zendy; Maria Docanto | Zendy; Nicole Ng | Zendy; Luciano G. Martelotto | Zendy; Christine Hauser | Zendy; Melissa C. Southey | Zendy; Arun Azad | Zendy; Tú NguyenDumont | Zendy

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Analytical Validation of an Error-Corrected Ultra-Sensitive Ctdna Next-Generation Sequencing Assay

Author(s) -

Heidi Fettke,

Jason A. Steen,

Edmond M. Kwan,

Patricia Bukczynska,

Shivakumar Keerthikumar,

David L. Goode,

Maria Docanto,

Nicole Ng,

Luciano G. Martelotto,

Christine Hauser,

Melissa C. Southey,

Arun Azad,

Tú NguyenDumont

Publication year - 2020

Publication title -

biotechniques

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 0.617

H-Index - 131

eISSN - 1940-9818

pISSN - 0736-6205

DOI - 10.2144/btn-2020-0045

Subject(s) - concordance , somatic cell , assay sensitivity , dna sequencing , biology , microbiology and biotechnology , dna , computational biology , chromatography , chemistry , genetics , medicine , gene , pathology , alternative medicine

Plasma circulating tumor DNA (ctDNA) analysis has emerged as a minimally invasive means to perform molecular tumor typing. Here we developed a custom ultra-sensitive ctDNA next-generation sequencing assay using molecular barcoding technology and off-the-shelf reagents combined with bioinformatics tools for enhanced ctDNA analysis. Assay performance was assessed via a spike-in experiment and the technique was applied to analyze 41 plasma samples from men with advanced prostate cancer. Orthogonal validation was performed using a commercial assay. Sensitivity and specificity of 93 and 99.5% were recorded for ultra-rare somatic variants (<1%), with high concordance observed between the in-house and commercial assays. The optimized protocol dramatically improved the efficiency of the assay and enabled the detection of low-frequency somatic variants from plasma cell-free DNA (cfDNA).

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