An electrochemical biosensor for the detection of Mycobacterium tuberculosis DNA from sputum and urine samples.
An electrochemical biosensor for the detection of Mycobacterium tuberculosis DNA from sputum and urine samples.
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DOI：
10.1371/journal.pone.0241067
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发表时间：
2020
期刊：
影响因子：
3.7
通讯作者：
Zimic M
中科院分区：
文献类型：
--
作者：
Ramos-Sono D;Laureano R;Rueda D;Gilman RH;La Rosa A;Ruiz J;León R;Sheen P;Zimic M
结核病（TB）是一个主要的全球性公共卫生问题，具有高死亡率和高发病率。在中低收入国家（LMIC），每年有大量需要结核病筛查的呼吸道症状病例，需要更准确、快速和负担得起的结核病诊断检测。痰涂片是LMIC的初始筛查试验，然而，其敏感性在痰菌负荷低的患者中是有限的。在目前可用的分子测试中观察到相同的限制。我们设计、标准化和评估了检测高度特异性DNA插入元件6110（IS 6110）的电化学生物传感器。将PCR扩增的DNA产物与固定的特异性DNA探针杂交在构建于FTO-玻璃上的工作电极的表面上，之后用Ag/AgCl参比电极和铂对电极进行循环伏安法。通过比率（杂交传感器的阴极峰值电流）/（非杂交传感器的阴极峰值电流）测量传感器的响应。我们测试了生物传感器，使用阳性杂交控制序列，从M.结核菌株和结核病人的痰，并从加标M.结核病DNA该生物传感器可有效检测M.结核病DNA的检测限为16 fM的痰样品和1 fM的加标尿样。低成本和相对较短的检测持续时间使其成为对抗结核病的重要结核病筛查工具。
Tuberculosis (TB) is a major global public health problem with high mortality and morbidity. In low-middle income countries (LMIC) a large number of respiratory symptomatic cases that require TB screening per year demands more accurate, fast and affordable testing for TB diagnostics. Sputum smear is the initial screening test in LMICs, however, its sensitivity is limited in patients with low sputum bacilli load. The same limitation is observed in the currently available molecular tests. We designed, standardized and evaluated an electrochemical biosensor that detects the highly specific DNA insertion element 6110 (IS6110). A PCR amplified DNA product is hybridized on the surface of the working electrode built on FTO-Glass with immobilized specific DNA probes, after which cyclic voltammetry is performed with an Ag/AgCl reference electrode and a platinum counter electrode. The response of the sensor was measured by the ratio (cathodic peak current of the hybridized sensor) / (cathodic peak current of the non-hybridized sensor). We tested the biosensor, using positive hybridization control sequences, genomic DNA extracted from M. tuberculosis strains and sputum of TB patients, and extracted DNA from the urine of healthy controls spiked with M. tuberculosis DNA. This biosensor was effective for the detection of M. tuberculosis DNA with a detection limit of 16 fM in sputum sample and 1 fM in spiked urine samples. The low cost and the relatively brief duration of the assay make this an important TB screening tool in the fight against tuberculosis.
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