Tuberculosis (TB) is one of the most lethal infectious diseases in the world. According to the annual report of the World Health Organization, TB caused an estimated 1.3 million deaths in 2017. The lack of effective diagnostic tool is one of the main reasons to cause this tragic result. Currently, there are several methods of diagnosing TB, including culture-based techniques, chest X-rays and CT scans, sputum smear microscopy, the enzyme-linked immunosorbent assay, and nucleic acid amplification tests. However, these methods often require complicated processing and costly detection tools to analyze the experimental results. Therefore, ongoing research has led to the development of various portable, hand-held, and point-of-care diagnostic methods that can be used in resource-limited settings.

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This study is highlighted as a Supplementary Cover of the ACS Sensors.

In this work, we investigate highly sensitive fluorescent Cu nanoparticles for use as rapid and specific nucleic acid amplification nanoprobes (NPs) for the diagnosis of tuberculosis. After applying polymerase chain reaction to a TB sample, we demonstrate that the presence of the targeted IS6110 DNA sequence of TB can be easily and directly detected through the in situ formation of DNA-templated fluorescent Cu-NPs and subsequently quantified using only a smartphone. Compared to traditional DNA analysis, this sensing platform does not require purification steps and eliminates the need for electrophoresis to confirm the PCR results. After optimization, this dsDNA-Cu-NPs-PCR method has the ability to analyze clinical TB nucleic acid samples at a detection limit of 5 fg/μL, and the fluorescent signal can be distinguished in only ~3 min after the DNA has been amplified. Moreover, with a combination of smartphone-assisted imaging analysis, we can further reduce the instrument size/cost and enhance the portability. In this manner, we are able to eliminate the need for a fluorescent spectrophotometer to measure the clinical sample. These results demonstrate this platform’s practical applicability, combining a smartphone and on-site analysis while retaining detection performance, making it suitable for clinical DNA applications in resource-limited regions of the world.

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The principle of using an AT-rich primer to simultaneously synthesize fluorescent dsDNA-Cu-NPs from a tuberculosis patient sample.

 

Contact: Prof. Chien-Fu Chen
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ACS Sensors
https://pubs.acs.org/doi/abs/10.1021/acssensors.9b01163

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