Assessment of real-time PCR method for detection of EGFR mutation using both supernatant and cell pellet of malignant pleural effusion samples from non-small-cell lung cancer patients

Saeam Shin, Juwon Kim, Yoonjung Kim, Sun Mi Cho, Kyung A. Lee

Research output: Contribution to journalArticlepeer-review

20 Citations (Scopus)

Abstract

EGFR mutation is an emerging biomarker for treatment selection in non-small-cell lung cancer (NSCLC) patients. However, optimal mutation detection is hindered by complications associated with the biopsy procedure, tumor heterogeneity and limited sensitivity of test methodology. In this study, we evaluated the diagnostic utility of real-time PCR using malignant pleural effusion samples. A total of 77 pleural fluid samples from 77 NSCLC patients were tested using the cobas EGFR mutation test (Roche Molecular Systems). Pleural fluid was centrifuged, and separated cell pellets and supernatants were tested in parallel. Results were compared with Sanger sequencing and/or peptide nucleic acid (PNA)-mediated PCR clamping of matched tumor tissue or pleural fluid samples. All samples showed valid real-time PCR results in one or more DNA samples extracted from cell pellets and supernatants. Compared with other molecular methods, the sensitivity of real-time PCR method was 100%. Concordance rate of real-time PCR and Sanger sequencing plus PNA-mediated PCR clamping was 98.7%. We have confirmed that real-time PCR using pleural fluid had a high concordance rate compared to conventional methods, with no failed samples. Our data demonstrated that the parallel real-time PCR testing using supernatant and cell pellet could offer reliable and robust surrogate strategy when tissue is not available.

Original languageEnglish
Pages (from-to)1962-1969
Number of pages8
JournalClinical Chemistry and Laboratory Medicine
Volume55
Issue number12
DOIs
Publication statusPublished - 2017 Oct 26

Bibliographical note

Funding Information:
Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission. Research funding: This study was supported by grants from the Korean Health Technology R&D Project of the Ministry of Health & Welfare (A120030), as well as the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning (NRF-2014R1A13053986). Employment or leadership: None declared. Honorarium: None declared. Competing interests: The funding organization(s) played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.

All Science Journal Classification (ASJC) codes

  • Clinical Biochemistry
  • Biochemistry, medical

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