Establishment of a new immunological method for direct detection of Mycobacterium in solution.
OBJECTIVE
Tuberculosis (TB) is a crucial health problem. Prevention of the disease requires rapid diagnosis. Rapid liquid culture systems, nucleic acid amplification tests, and high-performance liquid chromatography (HPLC) are among the rapid tests used for detecting Mycobacterium species. However, these tests are expensive and require extensive equipment and expertise, which is hardly affordable in resource-poor countries. Although direct microscopy is performed routinely as an initial step for detection of the bacteria, it is not sufficiently sensitive. As a result, we thought of establishing a low-cost immunological test that can potentially replace direct microscopy with higher sensitivity and specificity.


METHODS
The assay is based on pre-incubation of biotinylated rabbit antibody against Antigen 60 (A60) with a solution containing Bacillus Calmette-Guérin (BCG) or Mycobacterium tuberculosis (MTB) followed by incubation with a streptavidin-alkaline phosphatase (STA-ALP) conjugate. The test is devised in enzyme-linked immunosorbent assay (ELISA) and non-ELISA formats, therefore it does not require extensive facilities and expertise.


RESULTS
The ELISA format showed a 100-fold improvement in the lower detection limit of BCG compared with direct microscopy. With the non-ELISA formats, there was a 2- and 16-fold improvement for the cartridge assay and the microfuge tube assay, respectively.


CONCLUSIONS
In conclusion, we successfully detected BCG and MTB in solution using the new immunological method. Our results are very promising and the new immunological method could potentially replace direct microscopy with higher sensitivity and specificity.
Department of Medical Biotechnology and Cellular and Molecular Research Center, Iran University of Medical Sciences, Tehran, Iran.
Sepideh
S
Hamzehlou
Department of Medical Biotechnology and Cellular and Molecular Research Center, Iran University of Medical Sciences, Tehran, Iran. Electronic address: mfarajol@tums.ac.ir.
Mohammad M
MM
Farajollahi
D001431:Bacteriological Techniques / Q000379:methods*; D007118:Immunoassay / Q000379:methods*; D009161:Mycobacterium / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D014376:Tuberculosis / Q000175:diagnosis*
Avidin; BCG; Biotin; ELISA; Pre-incubation; Rapid immunodiagnostic test