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The accurate and rapid detection of Mycobacterium tuberculosis (M. tuberculosis) is essential for the effective treatment of tuberculosis. In this article, we propose an electrochemical sensor to detect M. tuberculosis reference strain H37Rv. The sensor contains an H37Rv aptamer and oligonucleotides modified with gold nanoparticles (AuNPs− DNA). An H37Rv aptamer screened by our laboratory was used as the recognition probe. The change in frequency shift mediated by AuNPs−DNA in the presence of H37Rv was detected using a multichannel series piezoelectric quartz crystal (MSPQC) system. Three oligonucleotides modified with gold nanoparticles were designed. These oligonucleotides contained 12, 12, and 13 bases that hybridized with the 37-nt H37Rv aptamer. H37Rv aptamer was immobilized on the gold electrode surface by Au−S bonds. A conductive-layer was then formed by sequential hybridization of the aptamer with the three designed AuNPs−DNAs. When H37Rv was present, it specifically bound to the aptamer, resulting in the detachment of AuNPs−DNA from the electrode. The conductive layer was thereby replaced by a nonconductive complex of aptamer and bacteria. These changes were monitored by the MSPQC system. The proposed sensor is rapid, specific and sensitive, the detection time was 2 h. The detection limit was 100 cfu/mL. This sensor would be of great benefit for the early clinical diagnosis of tuberculosis.

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Experimental Reagents and chemicalsA culture of attenuated M. tuberculosis (obtained from Dept. A piezoelectric immunosensor was developed for detecting Mycobacteria Tuberculosis (M. TB). Protein A was modified onto a silver-coated piezoelectric quartz-crystal sensor. Antibodies could be bond onto the crystal surface in an ordered orientation through protein A. An impendence analyzer was employed to record the admittance-frequency curve. The experimental results showed a successful and less-rigid bonding of protein A and antibodies. Compared with other traditional detection methods of TB, the method suggested in this paper was sensitive, selective and effective. The nonspecific response was limited by using control antibody-rabbit anti-honey bee venom (anti-HBV). A glycine-HCl buffer solution (pH = 2.4) was used to release antibodies from a crystal coated with protein A, and 0.5 mg/mL anti-TB was used to remove TB from a crystal that was bonded by an antibody. Good reusability was exhibited. Spike samples of sputum and saliva from normal people with and without adding M. tuberculosis were diagnosed using the proposed method. Good results were obtained.

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