Real-time PCR with internal amplification control for detecting tuberculosis: Method design and validation

E. Flores, J. C. RodrÍguez, E. Garcia-PachÓn, J. L. Soto, M. Ruiz, I. Escribano, G. Royo

Research output: Contribution to journalArticle

11 Scopus citations

Abstract

Flores E, Rodríguez JC, Garcia-Pachón E, Soto JL, Ruiz M, Escribano I, Royo G. Real-time PCR with internal amplification control for detecting tuberculosis: method design and validation. APMIS 2009; 117: 592-7. Real-time PCR has been a major development in the diagnosis of tuberculosis. However, most tests do not include an internal amplification control (IAC), which therefore limits it clinical application. In this study a new, easy to perform real-time PCR test with IAC was designed and validated in clinical samples. The primers amplified a 163-bp fragment of IS6110 of Mycobacterium tuberculosis and the IAC was designed with a fragment of a different microorganism (Chlamydia trachomatis). The interassay and intraassay variation of this test were very low (0.45-1.65% and 0.18-1.80%, respectively). The detection accuracy was validated in 50 samples (25 urine, 25 sputum) with different concentrations of M. tuberculosis, 18 clinical isolates of non-tuberculous mycobacteria and 148 samples with clinical suspicion of pulmonary tuberculosis. The specificity was 100%. The detection limit of this PCR test without IAC was approximately 15 bacteria and with IAC approximately 32 bacteria. This real-time PCR with IAC assay can improve the detection of M. tuberculosis and contribute to standardization of this diagnostic technique. © 2009 APMIS.
Original languageAmerican English
Pages (from-to)592-597
Number of pages6
JournalAPMIS
DOIs
StatePublished - 1 Aug 2009
Externally publishedYes

Fingerprint Dive into the research topics of 'Real-time PCR with internal amplification control for detecting tuberculosis: Method design and validation'. Together they form a unique fingerprint.

  • Cite this