SPECIFIC AND ACCURATE IDENTIFICATION OF MYCOBACTERIUM TUBERCULOSIS IN CLINICAL SAMPLES BY PCR-ELISA TECHNIQUE

Background and Aim:: Precise and timely diagnosis of tuberculosis is one of the important issues in the treatment management of this disease. Because of the slow growth of mycobacterium tuberculosis in the culture medium and the serious barriers of conventional methods such as microscopic testing, the urgent need for quick, sensitive and precise techniques to detect mycobacterium tuberculosis is felt. In this study, the aim was to detect M. tuberculosis, in specimens isolated from TB patients using PCR-ELISA with high sensitivity and specificity.Methods:: Using TUF7 and TUF4 specific primers, a 760 nucleotide fragment of the tuf gene from Mycobacterium tuberculosis was amplified by PCR. DIG-labeled amplicons were hybridized with a specific biotinylated oligonucleotide probe in solution phase and subsequently transferred to streptoavidin coated plates. The captured DNA were colorimetrically detected by the addition of anti-digoxigenin antibody HRP conjugate and 2,2-azino-di-(3-ethylbenzthiazolinsulfonate) substrate.Results:The results indicated a high sensitivity of PCR-ELISA for TB diagnosis. By employing this system, we could detect M. tuberculosis DNA as much as 20 pg in concentration and no interference was encountered in the amplification and detection of Mycobacterium tuberculosis in the presence of non-target DNA.Conclusion:The PCR-ELISA system offers several advantages in terms of sensitivity, rapidity and simplicity for detection M. tuberculosis in clinical specimens. Since there is a lot of problems in detecting a rapid and accurate TB disease, launching such a technique in laboratories can solve many of the leading problems in diagnosing and treating the disease.

Keywords:

Mycobacterium Tuberculosis , PCR-ELISA , tuf