Ultrafast, One-Step, Label-Based Biosensor Diagnosis Platform for the Detection of Mycobacterium tuberculosis in Clinical Applications.
ACS infectious diseases(2023)
Abstract
Tuberculosis (TB) is a chronic infectious disease caused by the etiological agent (MTB). Because the majority of TB patients come from poor economic backgrounds, the development of a simple, specific, low-cost, and highly sensitive detection method for the pathogen is extremely important for the prevention and treatment of this disease. In the current study, an efficient detection method for visual, rapid, and highly sensitive detection of MTB utilizing multiplex loop-mediated isothermal amplification combined with a label-based lateral flow immunoassay biosensor (mLAMP-LFIA) was developed. Three specific primer sets targeting the MTB genes 6110 and 64 were successfully designed and synthesized for the LAMP assay. The optimal reaction conditions for the mLAMP-LFIA assay were confirmed to be 67 °C for 40 min. The mLAMP amplicons were intuitively verified using the LFIA biosensor within 5 min. The entire process, including clinical sample processing, amplification reaction, and product verification, was completed within 80 min. The limit of detection of the mLAMP-LFIA assay established in the current study was 100 fg per reaction for the genomic DNA of MTB H37Rv. The analytical specificity of the mLAMP-LFIA assay was one hundred percent, and no cross-reactions with non-target strains were detected. Compared with the GeneXpert test, the sensitivity of mLAMP-LFIA for 148 clinical specimens was 100% (97/97), and the specificity was 98.04% (50/51) in the preliminary evaluation of the clinical application. Hence, the mLAMP-LFIA method, targeting the 6110 and 64 genes, is an ultrafast, one-step, low-cost, and highly sensitive detection method that could be used as a screening and/or diagnostic tool for MTB in the clinical setting, basic science laboratories, and especially in resource-poor regions.
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Key words
Mycobacterium tuberculosis,diagnosis,lateral flow immunoassay biosensor,loop-mediated isothermal amplification,tuberculosis
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