Droplet digital PCR-based EGFR mutation detection with an internal quality control index to determine the quality of DNA

• Published in: Scientific reports Vol. 8; no. 1; p. 543
• Main Authors: Kim, Sung-Su, Choi, Hyun-Jeung, Kim, Jin Ju, Kim, M. Sun, Lee, In-Seon, Byun, Bohyun, Jia, Lina, Oh, Myung Ryurl, Moon, Youngho, Park, Sarah, Choi, Joon-Seok, Chae, Seoung Wan, Nam, Byung-Ho, Kim, Jin-Soo, Kim, Jihun, Min, Byung Soh, Lee, Jae Seok, Won, Jae-Kyung, Cho, Soo Youn, Choi, Yoon-La, Shin, Young Kee
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 11.01.2018; Nature Publishing Group
• Subjects: 38/77; 692/53/2421; 692/700/139/1512; Biomarkers, Tumor - genetics; Biomarkers, Tumor - standards; Carcinoma, Non-Small-Cell Lung - diagnosis; Carcinoma, Non-Small-Cell Lung - genetics; Deoxyribonucleic acid; DNA; DNA - chemistry; DNA - genetics; DNA - standards; Epidermal growth factor receptors; ErbB Receptors - genetics; Genetic testing; Genomes; Humanities and Social Sciences; Humans; Lung cancer; Lung Neoplasms - diagnosis; Lung Neoplasms - genetics; Molecular Diagnostic Techniques - methods; Molecular Diagnostic Techniques - standards; multidisciplinary; Mutation; Non-small cell lung carcinoma; Paraffin; Polymerase chain reaction; Polymerase Chain Reaction - methods; Polymerase Chain Reaction - standards; Quality control; Reference Standards; Science; Science (multidisciplinary)
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-017-18642-x

In clinical translational research and molecular in vitro diagnostics, a major challenge in the detection of genetic mutations is overcoming artefactual results caused by the low-quality of formalin-fixed paraffin-embedded tissue (FFPET)-derived DNA (FFPET-DNA). Here, we propose the use of an ‘internal quality control (iQC) index’ as a criterion for judging the minimum quality of DNA for PCR-based analyses. In a pre-clinical study comparing the results from droplet digital PCR-based EGFR mutation test (ddEGFR test) and qPCR-based EGFR mutation test (cobas EGFR test), iQC index ≥ 0.5 (iQC copies ≥ 500, using 3.3 ng of FFPET-DNA [1,000 genome equivalents]) was established, indicating that more than half of the input DNA was amplifiable. Using this criterion, we conducted a retrospective comparative clinical study of the ddEGFR and cobas EGFR tests for the detection of EGFR mutations in non-small cell lung cancer (NSCLC) FFPET-DNA samples. Compared with the cobas EGFR test, the ddEGFR test exhibited superior analytical performance and equivalent or higher clinical performance. Furthermore, iQC index is a reliable indicator of the quality of FFPET-DNA and could be used to prevent incorrect diagnoses arising from low-quality samples.