Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2
The COVID-19 pandemic has presented a significant challenge to the world's public health and led to over 6.9 million deaths reported to date. A rapid, sensitive, and cost-effective point-of-care virus detection device is essential for the control and surveillance of the contagious severe acute...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-09-01
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| Series: | Biosensors and Bioelectronics: X |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S259013702300078X |
| _version_ | 1797733874175836160 |
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| author | Xiaochen Qin Ratul Paul Yuyuan Zhou Yue Wu Xuanhong Cheng Yaling Liu |
| author_facet | Xiaochen Qin Ratul Paul Yuyuan Zhou Yue Wu Xuanhong Cheng Yaling Liu |
| author_sort | Xiaochen Qin |
| collection | DOAJ |
| description | The COVID-19 pandemic has presented a significant challenge to the world's public health and led to over 6.9 million deaths reported to date. A rapid, sensitive, and cost-effective point-of-care virus detection device is essential for the control and surveillance of the contagious severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic. The study presented here aimed to demonstrate a solid-phase isothermal recombinase polymerase amplification coupled CRISPR-based (spRPA-CRISPR) assay for on-chip multiplexed, sensitive and visual COVID-19 DNA detection. The assay targets the SARS-CoV-2 structure protein encoded genomes and can simultaneously detect two specific genes without cross-interaction. The amplified target sequences were immobilized on the one-pot device surface and detected using the mixed Cas12a-crRNA collateral cleavage of reporter-released fluorescent signal when specific genes were recognized. The endpoint signal can be directly visualized for rapid detection of COVID-19. The system was tested with samples of a broad range of concentrations (20 to 2 × 104 copies) and showed analytical sensitivity down to 20 copies per microliter. Furthermore, a low-cost blue LED flashlight (∼$12) was used to provide a visible SARS-CoV-2 detection signal of the spRPA-CRISPR assay which could be purchased online easily. Thus, our platform provides a sensitive and easy-to-read multiplexed gene detection method that can specifically identify low concentration genes. |
| first_indexed | 2024-03-12T12:34:49Z |
| format | Article |
| id | doaj.art-13db827e75c64943828bfca13156ae59 |
| institution | Directory Open Access Journal |
| issn | 2590-1370 |
| language | English |
| last_indexed | 2024-03-12T12:34:49Z |
| publishDate | 2023-09-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Biosensors and Bioelectronics: X |
| spelling | doaj.art-13db827e75c64943828bfca13156ae592023-08-29T04:17:56ZengElsevierBiosensors and Bioelectronics: X2590-13702023-09-0114100381Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2Xiaochen Qin0Ratul Paul1Yuyuan Zhou2Yue Wu3Xuanhong Cheng4Yaling Liu5Department of Bioengineering, Lehigh University, Bethlehem, PA, 18015, USADepartment of Mechanical Engineering & Mechanics, Lehigh University, Bethlehem, PA, 18015, USADepartment of Bioengineering, Lehigh University, Bethlehem, PA, 18015, USADepartment of Bioengineering, Lehigh University, Bethlehem, PA, 18015, USADepartment of Bioengineering, Lehigh University, Bethlehem, PA, 18015, USA; Department of Materials Science and Engineering, Lehigh University, Bethlehem, PA, 18015, USADepartment of Bioengineering, Lehigh University, Bethlehem, PA, 18015, USA; Department of Mechanical Engineering & Mechanics, Lehigh University, Bethlehem, PA, 18015, USA; Corresponding author. Department of Bioengineering, Lehigh University, Bethlehem, PA, 18015, USA.The COVID-19 pandemic has presented a significant challenge to the world's public health and led to over 6.9 million deaths reported to date. A rapid, sensitive, and cost-effective point-of-care virus detection device is essential for the control and surveillance of the contagious severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic. The study presented here aimed to demonstrate a solid-phase isothermal recombinase polymerase amplification coupled CRISPR-based (spRPA-CRISPR) assay for on-chip multiplexed, sensitive and visual COVID-19 DNA detection. The assay targets the SARS-CoV-2 structure protein encoded genomes and can simultaneously detect two specific genes without cross-interaction. The amplified target sequences were immobilized on the one-pot device surface and detected using the mixed Cas12a-crRNA collateral cleavage of reporter-released fluorescent signal when specific genes were recognized. The endpoint signal can be directly visualized for rapid detection of COVID-19. The system was tested with samples of a broad range of concentrations (20 to 2 × 104 copies) and showed analytical sensitivity down to 20 copies per microliter. Furthermore, a low-cost blue LED flashlight (∼$12) was used to provide a visible SARS-CoV-2 detection signal of the spRPA-CRISPR assay which could be purchased online easily. Thus, our platform provides a sensitive and easy-to-read multiplexed gene detection method that can specifically identify low concentration genes.http://www.sciencedirect.com/science/article/pii/S259013702300078XCOVID-19CRISPR-Based assayMultiplexed spRPA-CRISPR assayDNA sensorPoint-of-careVisual detection |
| spellingShingle | Xiaochen Qin Ratul Paul Yuyuan Zhou Yue Wu Xuanhong Cheng Yaling Liu Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2 Biosensors and Bioelectronics: X COVID-19 CRISPR-Based assay Multiplexed spRPA-CRISPR assay DNA sensor Point-of-care Visual detection |
| title | Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2 |
| title_full | Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2 |
| title_fullStr | Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2 |
| title_full_unstemmed | Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2 |
| title_short | Multiplex solid-phase RPA coupled CRISPR-based visual detection of SARS-CoV-2 |
| title_sort | multiplex solid phase rpa coupled crispr based visual detection of sars cov 2 |
| topic | COVID-19 CRISPR-Based assay Multiplexed spRPA-CRISPR assay DNA sensor Point-of-care Visual detection |
| url | http://www.sciencedirect.com/science/article/pii/S259013702300078X |
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