Solid-Phase Microextraction of DNA from Mycobacteria in Artificial Sputum Samples To Enable Visual Detection Using Isothermal Amplification
Point-of-care (POC) technologies for the detection of pathogens in clinical samples are highly valued due to their speed, ease of use, and cost-effectiveness. Furthermore, they are ideally suited for resource-limited settings where expensive and sophisticated laboratory equipment may not be readily...
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| Veröffentlicht in: | Analytical chemistry (Washington) 2018-06, Vol.90 (11), p.6922-6928 |
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| creator | Varona, Marcelino Ding, Xiong Clark, Kevin D Anderson, Jared L |
| description | Point-of-care (POC) technologies for the detection of pathogens in clinical samples are highly valued due to their speed, ease of use, and cost-effectiveness. Furthermore, they are ideally suited for resource-limited settings where expensive and sophisticated laboratory equipment may not be readily available. In this study, a rapid method based on solid-phase microextraction (SPME) of mycobacterial DNA with subsequent isothermal amplification and visual detection was developed. Direct coupling of the SPME desorption solution (1 M NaCl) to the isothermal reaction system was achieved to circumvent dilution steps and improve detection limits. Using this method, DNA was preconcentrated from lysed mycobacteria in just 2 min, subjected to isothermal multiple-self-matching-initiated amplification (IMSA), and the amplicons were detected visually. With a total analysis times of less than 2 h, the optimized method was capable of extracting and visually detecting mycobacterial DNA from artificial sputum samples containing clinically relevant concentrations of mycobacteria (107 colony forming units/mL), demonstrating its potential for future POC applications. |
| doi_str_mv | 10.1021/acs.analchem.8b01160 |
| format | Article |
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With a total analysis times of less than 2 h, the optimized method was capable of extracting and visually detecting mycobacterial DNA from artificial sputum samples containing clinically relevant concentrations of mycobacteria (107 colony forming units/mL), demonstrating its potential for future POC applications.</description><subject>Amplification</subject><subject>Analytical chemistry</subject><subject>Bacteria</subject><subject>Chemistry</subject><subject>Deoxyribonucleic acid</subject><subject>Dilution</subject><subject>DNA</subject><subject>Extraction processes</subject><subject>Gene amplification</subject><subject>Laboratory equipment</subject><subject>Sodium chloride</subject><subject>Solid phases</subject><subject>Solid state physics</subject><subject>Visual perception</subject><issn>0003-2700</issn><issn>1520-6882</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2018</creationdate><recordtype>article</recordtype><recordid>eNp9kc1u2zAQhIkiReOkfYOgIJBLL3KXlERaRyP_QH4KOOlVWNKrmIEkuqQENM-Qlw4NOznkkAsJcL-ZBWcYOxIwFSDFb7Rxij22dkXddGZACAVf2ESUEjI1m8k9NgGAPJMaYJ8dxPgEiQGhvrF9WelSK6Em7GXhW7fM_qwwEr9xNnj6PwS0g_M99w0_vZ3zJviO3zxbb9I7BYfc9XweBtc467Dli_U4jB1fYLduKfJ7z896NC3xvy6OaX5KA20NH6LrH_lV9MOKQpdG8yTZ2OBm_J19bbCN9GN3H7KH87P7k8vs-u7i6mR-nWGu1ZBRDkoUJVSmKEuBjTQGG1uaAitTLi2gksoSpc-DtVU68qKoFFZazkhqLfJD9mvruw7-30hxqDsXLbUt9uTHWEvIK1mplFZCjz-gT34MKfVECSiETonqRBVbKsUXY6CmXgfXYXiuBdSbsupUVv1WVr0rK8l-7sxH09HyXfTWTgJgC2zk74s_9XwFoeSkRQ</recordid><startdate>20180605</startdate><enddate>20180605</enddate><creator>Varona, Marcelino</creator><creator>Ding, Xiong</creator><creator>Clark, Kevin D</creator><creator>Anderson, Jared L</creator><general>American Chemical Society</general><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QF</scope><scope>7QO</scope><scope>7QQ</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7TA</scope><scope>7TB</scope><scope>7TM</scope><scope>7U5</scope><scope>7U7</scope><scope>7U9</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>H94</scope><scope>JG9</scope><scope>JQ2</scope><scope>KR7</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0003-0437-0589</orcidid><orcidid>https://orcid.org/0000-0001-6915-8752</orcidid></search><sort><creationdate>20180605</creationdate><title>Solid-Phase Microextraction of DNA from Mycobacteria in Artificial Sputum Samples To Enable Visual Detection Using Isothermal Amplification</title><author>Varona, Marcelino ; 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| subjects | Amplification Analytical chemistry Bacteria Chemistry Deoxyribonucleic acid Dilution DNA Extraction processes Gene amplification Laboratory equipment Sodium chloride Solid phases Solid state physics Visual perception |
| title | Solid-Phase Microextraction of DNA from Mycobacteria in Artificial Sputum Samples To Enable Visual Detection Using Isothermal Amplification |
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