Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus
A two-stage isothermal amplification method, which consists of a first-stage basic recombinase polymerase amplification (RPA) and a second-stage fluorescence loop-mediated isothermal amplification (LAMP), as well as a microfluidic-chip-based portable system, were developed in this study; these enabl...
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| Format: | Article |
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MDPI AG
2021-12-01
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| Series: | Micromachines |
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| Online Access: | https://www.mdpi.com/2072-666X/12/12/1582 |
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| author | Qin Huang Xiaohui Shan Ranran Cao Xiangyu Jin Xue Lin Qiurong He Yulei Zhu Rongxin Fu Wenli Du Wenqi Lv Ying Xia Guoliang Huang |
| author_facet | Qin Huang Xiaohui Shan Ranran Cao Xiangyu Jin Xue Lin Qiurong He Yulei Zhu Rongxin Fu Wenli Du Wenqi Lv Ying Xia Guoliang Huang |
| author_sort | Qin Huang |
| collection | DOAJ |
| description | A two-stage isothermal amplification method, which consists of a first-stage basic recombinase polymerase amplification (RPA) and a second-stage fluorescence loop-mediated isothermal amplification (LAMP), as well as a microfluidic-chip-based portable system, were developed in this study; these enabled parallel detection of multiplex targets in real time in around one hour, with high sensitivity and specificity, without cross-contamination. The consumption of the sample and the reagent was 2.1 μL and 10.6 μL per reaction for RPA and LAMP, respectively. The lowest detection limit (LOD) was about 10 copies. The clinical amplification of about 40 nasopharyngeal swab samples, containing 17 SARS-CoV-2 (severe acute respiratory syndrome coronavirus) and 23 measles viruses (MV), were parallel tested by using the microfluidic chip. Both clinical specificity and sensitivity were 100% for MV, and the clinical specificity and sensitivity were 94.12% and 95.83% for SARS-CoV-2, respectively. This two-stage isothermal amplification method based on the microfluidic chip format offers a convenient, clinically parallel molecular diagnostic method, which can identify different nucleic acid samples simultaneously and in a timely manner, and with a low cost of the reaction reagent. It is especially suitable for resource-limited areas and point-of-care testing (POCT). |
| first_indexed | 2024-03-10T03:32:35Z |
| format | Article |
| id | doaj.art-b0e2fc7b47764f978dd963a471661fe6 |
| institution | Directory Open Access Journal |
| issn | 2072-666X |
| language | English |
| last_indexed | 2024-03-10T03:32:35Z |
| publishDate | 2021-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Micromachines |
| spelling | doaj.art-b0e2fc7b47764f978dd963a471661fe62023-11-23T09:37:21ZengMDPI AGMicromachines2072-666X2021-12-011212158210.3390/mi12121582Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles VirusQin Huang0Xiaohui Shan1Ranran Cao2Xiangyu Jin3Xue Lin4Qiurong He5Yulei Zhu6Rongxin Fu7Wenli Du8Wenqi Lv9Ying Xia10Guoliang Huang11Department of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaSichuan Center for Disease Control and Prevention, Chengdu 610041, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaWest China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu 610041, ChinaWest China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu 610041, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaWest China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu 610041, ChinaDepartment of Biomedical Engineering, School of Medicine, Tsinghua University, Beijing 100084, ChinaA two-stage isothermal amplification method, which consists of a first-stage basic recombinase polymerase amplification (RPA) and a second-stage fluorescence loop-mediated isothermal amplification (LAMP), as well as a microfluidic-chip-based portable system, were developed in this study; these enabled parallel detection of multiplex targets in real time in around one hour, with high sensitivity and specificity, without cross-contamination. The consumption of the sample and the reagent was 2.1 μL and 10.6 μL per reaction for RPA and LAMP, respectively. The lowest detection limit (LOD) was about 10 copies. The clinical amplification of about 40 nasopharyngeal swab samples, containing 17 SARS-CoV-2 (severe acute respiratory syndrome coronavirus) and 23 measles viruses (MV), were parallel tested by using the microfluidic chip. Both clinical specificity and sensitivity were 100% for MV, and the clinical specificity and sensitivity were 94.12% and 95.83% for SARS-CoV-2, respectively. This two-stage isothermal amplification method based on the microfluidic chip format offers a convenient, clinically parallel molecular diagnostic method, which can identify different nucleic acid samples simultaneously and in a timely manner, and with a low cost of the reaction reagent. It is especially suitable for resource-limited areas and point-of-care testing (POCT).https://www.mdpi.com/2072-666X/12/12/1582two-stage isothermal amplificationmicrofluidic chipparallel molecular diagnosticSARS-CoV-2measles virus |
| spellingShingle | Qin Huang Xiaohui Shan Ranran Cao Xiangyu Jin Xue Lin Qiurong He Yulei Zhu Rongxin Fu Wenli Du Wenqi Lv Ying Xia Guoliang Huang Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus Micromachines two-stage isothermal amplification microfluidic chip parallel molecular diagnostic SARS-CoV-2 measles virus |
| title | Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus |
| title_full | Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus |
| title_fullStr | Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus |
| title_full_unstemmed | Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus |
| title_short | Microfluidic Chip with Two-Stage Isothermal Amplification Method for Highly Sensitive Parallel Detection of SARS-CoV-2 and Measles Virus |
| title_sort | microfluidic chip with two stage isothermal amplification method for highly sensitive parallel detection of sars cov 2 and measles virus |
| topic | two-stage isothermal amplification microfluidic chip parallel molecular diagnostic SARS-CoV-2 measles virus |
| url | https://www.mdpi.com/2072-666X/12/12/1582 |
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