en میکروب شناسی Microbiology مقالات اصلی Original Article <div style="text-align: justify;"><strong><em>Background:</em></strong> The line probe assay (LPA) is one of the most accurate diagnostic tools for detection of different <em>Mycobacterium</em> species. Several commercial kits based on the LPA for detection of <em>Mycobacterium</em> species are currently available. Because of their high cost, especially for underdeveloped and developing countries, and the discrepancy of non-tuberculous mycobacteria (NTM) prevalence across geographic regions, it would be reasonable to consider the development of an in-house LPA. The aim of this study was to develop an LPA to detect and differentiate mycobacterial species and to evaluate the usefulness of PCR-LPA for direct application on clinical samples.<br> <br> <strong><em>Methods:</em></strong> One pair of biotinylated primers and 15 designed DNA oligonucleotide probes were used based on multiple aligned internal transcribed spacer (ITS) sequences. Specific binding of the PCR-amplified products to the probes immobilized on nitrocellulose membrane strips was evaluated by the hybridization method. Experiments were performed three times on separate days to evaluate the assay&rsquo;s repeatability. The PCR-LPA was evaluated directly on nine clinical samples and their cultivated isolates.<br> <br> <strong><em>Results:</em></strong> All 15 probes used in this study hybridized specifically to ITS sequences of the corresponding standard species. Results were reproducible for all the strains on different days. <em>Mycobacterium</em> species of the nine clinical specimens and their cultivated isolates were correctly identified by PCR-LPA and confirmed by sequencing.<br> <br> <strong><em>Conclusions:</em></strong> In this study, we describe a PCR-LPA that is readily applicable in the clinical laboratory. The assay is fast, cost-effective, highly specific, and requires no radioactive materials.<br> &nbsp;</div> Diagnosis, Line Probe Assay (LPA), Mycobacterium Infection, Tuberculosis. 383 393 http://rbmb.net/browse.php?a_code=A-10-334-2&slc_lang=en&sid=1 Reza kamali Kakhki reza.kamali99@yahoo.com 100319475328460017415 100319475328460017415 No Antimicrobial Resistance Research Center, Medical School, Mashhad University of Medical Sciences, Mashhad, Iran. Ehsan Aryan aryanE@mums.ac.ir 100319475328460017416 100319475328460017416 No Antimicrobial Resistance Research Center, Medical School, Mashhad University of Medical Sciences, Mashhad, Iran. Zahra Meshkat meshkatz@mums.ac.ir 100319475328460017417 100319475328460017417 No Antimicrobial Resistance Research Center, Medical School, Mashhad University of Medical Sciences, Mashhad, Iran. Mojtaba Sankian sankianm@mums.ac.ir 100319475328460017418 100319475328460017418 Yes Immunobiochemistry Laboratory, Immunology Research Center, Medical School, Mashhad University of Medical Sciences, Mashhad, Iran.