Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker
We introduce a digital microfluidics (DMF) platform specifically designed to perform a loop-mediated isothermal amplification (LAMP) of DNA and applied it to a real-time amplification to monitor a cancer biomarker, <i>c-Myc</i> (associated to 40% of all human tumors), using fluorescence...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2022-03-01
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| Series: | Biosensors |
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| Online Access: | https://www.mdpi.com/2079-6374/12/4/201 |
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| author | Beatriz Jorge Coelho Bruno Veigas Luís Bettencourt Hugo Águas Elvira Fortunato Rodrigo Martins Pedro V. Baptista Rui Igreja |
| author_facet | Beatriz Jorge Coelho Bruno Veigas Luís Bettencourt Hugo Águas Elvira Fortunato Rodrigo Martins Pedro V. Baptista Rui Igreja |
| author_sort | Beatriz Jorge Coelho |
| collection | DOAJ |
| description | We introduce a digital microfluidics (DMF) platform specifically designed to perform a loop-mediated isothermal amplification (LAMP) of DNA and applied it to a real-time amplification to monitor a cancer biomarker, <i>c-Myc</i> (associated to 40% of all human tumors), using fluorescence microscopy. We demonstrate the full manipulation of the sample and reagents on the DMF platform, resulting in the successful amplification of 90 pg of the target DNA (0.5 ng/µL) in less than one hour. Furthermore, we test the efficiency of an innovative mixing strategy in DMF by employing two mixing methodologies onto the DMF droplets—low frequency AC (alternating current) actuation as well as back-and-forth droplet motion—which allows for improved fluorescence readouts. Fluorophore bleaching effects are minimized through on-chip sample partitioning by DMF processes and sequential droplet irradiation. Finally, LAMP reactions require only 2 µL volume droplets, which represents a 10-fold volume reduction in comparison to benchtop LAMP. |
| first_indexed | 2024-03-09T11:05:57Z |
| format | Article |
| id | doaj.art-39d7d8592c37484fbe99502e763447c7 |
| institution | Directory Open Access Journal |
| issn | 2079-6374 |
| language | English |
| last_indexed | 2024-03-09T11:05:57Z |
| publishDate | 2022-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biosensors |
| spelling | doaj.art-39d7d8592c37484fbe99502e763447c72023-12-01T00:57:46ZengMDPI AGBiosensors2079-63742022-03-0112420110.3390/bios12040201Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer BiomarkerBeatriz Jorge Coelho0Bruno Veigas1Luís Bettencourt2Hugo Águas3Elvira Fortunato4Rodrigo Martins5Pedro V. Baptista6Rui Igreja7Department of Materials Science, School of Science and Technology, NOVA University of Lisbon and CEMOP/UNINOVA, Campus de Caparica, 2829-516 Caparica, PortugalAlmaScience, Campus da Caparica, 2829-519 Caparica, PortugalDepartment of Materials Science, School of Science and Technology, NOVA University of Lisbon and CEMOP/UNINOVA, Campus de Caparica, 2829-516 Caparica, PortugalDepartment of Materials Science, School of Science and Technology, NOVA University of Lisbon and CEMOP/UNINOVA, Campus de Caparica, 2829-516 Caparica, PortugalDepartment of Materials Science, School of Science and Technology, NOVA University of Lisbon and CEMOP/UNINOVA, Campus de Caparica, 2829-516 Caparica, PortugalDepartment of Materials Science, School of Science and Technology, NOVA University of Lisbon and CEMOP/UNINOVA, Campus de Caparica, 2829-516 Caparica, PortugalUCIBIO, I4HB, Life Sciences Department, School of Science and Technology, NOVA University of Lisbon, Campus de Caparica, 2829-516 Caparica, PortugalDepartment of Materials Science, School of Science and Technology, NOVA University of Lisbon and CEMOP/UNINOVA, Campus de Caparica, 2829-516 Caparica, PortugalWe introduce a digital microfluidics (DMF) platform specifically designed to perform a loop-mediated isothermal amplification (LAMP) of DNA and applied it to a real-time amplification to monitor a cancer biomarker, <i>c-Myc</i> (associated to 40% of all human tumors), using fluorescence microscopy. We demonstrate the full manipulation of the sample and reagents on the DMF platform, resulting in the successful amplification of 90 pg of the target DNA (0.5 ng/µL) in less than one hour. Furthermore, we test the efficiency of an innovative mixing strategy in DMF by employing two mixing methodologies onto the DMF droplets—low frequency AC (alternating current) actuation as well as back-and-forth droplet motion—which allows for improved fluorescence readouts. Fluorophore bleaching effects are minimized through on-chip sample partitioning by DMF processes and sequential droplet irradiation. Finally, LAMP reactions require only 2 µL volume droplets, which represents a 10-fold volume reduction in comparison to benchtop LAMP.https://www.mdpi.com/2079-6374/12/4/201digital microfluidicsloop-mediated isothermal amplificationreal-time nucleic acid amplification monitoringfluorescence detectioncancer biomarker |
| spellingShingle | Beatriz Jorge Coelho Bruno Veigas Luís Bettencourt Hugo Águas Elvira Fortunato Rodrigo Martins Pedro V. Baptista Rui Igreja Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker Biosensors digital microfluidics loop-mediated isothermal amplification real-time nucleic acid amplification monitoring fluorescence detection cancer biomarker |
| title | Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker |
| title_full | Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker |
| title_fullStr | Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker |
| title_full_unstemmed | Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker |
| title_short | Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker |
| title_sort | digital microfluidics powered real time monitoring of isothermal dna amplification of cancer biomarker |
| topic | digital microfluidics loop-mediated isothermal amplification real-time nucleic acid amplification monitoring fluorescence detection cancer biomarker |
| url | https://www.mdpi.com/2079-6374/12/4/201 |
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