Digital droplet RT-LAMP increases speed of SARS-CoV-2 viral RNA detection

被引：2
作者：
Yuan, Yuan [1 ,2 ]
Ellis, Perry [2 ]
Tao, Ye [2 ]
Bikos, Dimitri A. [3 ,4 ]
Loveday, Emma K. [3 ,4 ]
Thomas, Mallory M. [3 ,4 ]
Wilking, James N. [3 ,4 ,5 ]
Chang, Connie B. [3 ,4 ,5 ]
Ye, Fangfu [1 ,6 ,11 ]
Weitz, David A. [2 ,7 ,8 ,9 ,10 ]
机构：
[1] Univ Chinese Acad Sci, Wenzhou Inst, Oujiang Lab, Zhejiang Lab Regenerat Med Vis & Brain Hlth, Wenzhou, Zhejiang, Peoples R China
[2] Harvard Univ, John A Paulson Sch Engn & Appl Sci, Cambridge, MA 02138 USA
[3] Montana State Univ, Dept Chem & Biol Engn, Bozeman, MT USA
[4] Montana State Univ, Ctr Biofilm Engn, Bozeman, MT USA
[5] Mayo Clin, Dept Physiol & Biomed Engn, Rochester, MN USA
[6] Chinese Acad Sci, Beijing Natl Lab Condensed Matter Phys, Inst Phys, Beijing, Peoples R China
[7] Harvard Univ, Dept Phys, Cambridge, MA 02138 USA
[8] Harvard Univ, Wyss Inst Biol Inspired Engn, Boston, MA USA
[9] Harvard Univ, Dept Phys, Phys & Appl Phys, Pierce Hall 231,29 Oxford St, Cambridge, MA 02138 USA
[10] Harvard Univ, SEAS, Pierce Hall 231,29 Oxford St, Cambridge, MA 02138 USA
[11] Chinese Acad Sci, Inst Phys, 8 Zhongguancun 3rd South St, Beijing 100190, Peoples R China
来源：
SMART MEDICINE | 2024年 / 3卷 / 02期
基金：
美国国家卫生研究院; 中国国家自然科学基金;
关键词：
ddRT-LAMP; droplet microfluidics; droplet size; RNA rapid detection; SARS-CoV-2; virus; ACCURACY; PCR;
D O I：
10.1002/SMMD.20240008
中图分类号：
R318 [生物医学工程];
学科分类号：
0831 ;
摘要：
Nucleic acid amplification testing (NAAT) remains one of the most reliable methods for pathogen identification. However, conventional bulk NAATs may not be sufficiently fast or sensitive enough for the detection of clinically-relevant pathogens in point-of-care testing. Here, we have developed a digital droplet RT-LAMP (ddRT-LAMP) assay that rapidly and quantitatively detects the SARS-CoV-2 viral E gene in microfluidic drops. Droplet partitioning using ddRT-LAMP significantly accelerates detection times across a wide range of template concentrations compared to bulk RT-LAMP assays. We discover that a reduction in droplet diameter decreases assay times up to a certain size, upon which surface adsorption of the RT-LAMP polymerase reduces reaction efficiency. Optimization of drop size and polymerase concentration enables rapid, sensitive, and quantitative detection of the SARS-CoV-2 E gene in only 8 min. These results highlight the potential of ddRT-LAMP assays as an excellent platform for quantitative point-of-care testing. We have developed a digital droplet RT-LAMP (ddRT-LAMP) assay that rapidly and quantitatively detects the SARS-CoV-2 viral E gene in microfluidic drops. Droplet partitioning using ddRT-LAMP significantly accelerates detection times across a wide range of template concentrations compared to bulk RT-LAMP assays. image
引用
收藏
页数：9
相关论文
共 50 条
[11]   RT-LAMP Multicenter Study for SARS-CoV-2 Genome Molecular Detection in Brazilian Swab and Saliva Samples [J].
da Costa, Vanessa Duarte ;
Santos, Alanna Calheiros ;
da Silva, Lucas Lima ;
Wiggers, Wilian Jean ;
Ivantes, Claudia Alexandra Pontes ;
Lima, Danielle Malta ;
Colares, Jeova Keny Baima ;
Dallacqua, Deusilene Souza Vieira ;
Motta-Castro, Ana Rita Coimbra ;
Davila, Alberto Martin Rivera ;
Teles, Sheila Araujo ;
Carneiro, Megmar Aparecida dos Santos ;
Caetano, Karlla Antonieta Amorim ;
Anunciacao, Fernando Antonio Costa ;
de Paula, Vanessa Salete ;
Villar, Livia Melo .
DIAGNOSTICS, 2023, 13 (02)
[12]   Clinical Validation and Evaluation of a Colorimetric SARS-CoV-2 RT-LAMP Assay Against RT-PCR [J].
Erdem, Murat ;
Andac-Ozketen, Ayse ;
Ozketen, Ahmet Caglar ;
Karahan, Gizem ;
Tozluyurt, Abdullah ;
Palaz, Fahreddin ;
Alp, Alpaslan ;
Unal, Serhat .
INFECTIOUS DISEASES AND CLINICAL MICROBIOLOGY, 2023, 5 (02) :136-143
[13]   Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-Care Platform [J].
Donia, Ahmed ;
Shahid, Muhammad Furqan ;
Sammer-Ul Hassan ;
Shahid, Ramla ;
Ahmad, Aftab ;
Javed, Aneela ;
Nawaz, Muhammad ;
Yaqub, Tahir ;
Bokhari, Habib .
FOOD AND ENVIRONMENTAL VIROLOGY, 2022, 14 (4) :364-373
[14]   Starting from scratch: Step-by-step development of diagnostic tests for SARS-CoV-2 detection by RT-LAMP [J].
Tapia-Sidas, Diana Angelica ;
Vargas-Hernandez, Brenda Yazmin ;
Ramirez-Pool, Jose Abrahan ;
Nunez-Munoz, Leandro Alberto ;
Calderon-Perez, Berenice ;
Gonzalez-Gonzalez, Rogelio ;
Brieba, Luis Gabriel ;
Lira-Carmona, Rosalia ;
Ferat-Osorio, Eduardo ;
Lopez-Macias, Constantino ;
Ruiz-Medrano, Roberto ;
Xoconostle-Cazares, Beatriz .
PLOS ONE, 2023, 18 (01)
[15]   Multiplex, Real-Time, Point-of-care RT-LAMP for SARS-CoV-2 Detection Using the HFman Probe [J].
Dong, Yajuan ;
Zhao, Yongjuan ;
Li, Shenwei ;
Wan, Zhenzhou ;
Lu, Renfei ;
Yang, Xianguang ;
Yu, Guoying ;
Reboud, Julien ;
Cooper, Jonathan M. ;
Tian, Zhengan ;
Zhang, Chiyu .
ACS SENSORS, 2022, 7 (03) :730-739
[16]   Development of an optimized colorimetric RT-LAMP for SARS-CoV-2 assay with enhanced procedure controls for remote diagnostics [J].
Raddatz, Bruna Winkert ;
Kim, Edson Yu Sin ;
Imamura, Louise Matie ;
Steil, Gisleine Jarenko ;
Santiago, Erika Bergamo ;
Soares, Santiago Pedro Timm ;
Ribeiro, Victor Henrique Alves ;
de Almeida, Bernardo Montesanti Machado ;
Rogal Jr, Sergio Renato ;
Figueredo, Marcus Vinicius Mazega .
SCIENTIFIC REPORTS, 2022, 12 (01)
[17]   Detection of SARS-CoV-2 RNA by direct RT-qPCR on nasopharyngeal specimens without extraction of viral RNA [J].
Hasan, Mohammad Rubayet ;
Mirza, Faheem ;
Al-Hail, Hamad ;
Sundararaju, Sathyavathi ;
Xaba, Thabisile ;
Iqbal, Muhammad ;
Alhussain, Hashim ;
Yassine, Hadi Mohamad ;
Perez-Lopez, Andres ;
Tang, Patrick .
PLOS ONE, 2020, 15 (07)
[18]   Comparison of diagnostic performance of RT-qPCR, RT-LAMP and IgM/IgG rapid tests for detection of SARS-CoV-2 among healthcare workers in Brazil [J].
Bernardes, Wilma Patricia de Oliveira Santos ;
Santos, Thais Garcia ;
Fernandes, Nubia Monteiro Goncalves Soares ;
Silva, Thais Barbara de Souza ;
Westin, Mateus ;
Simoes, Taynana Cesar ;
Fernandes e Silva, Eduardo ;
Alves, Breno Magalhaes ;
Molina, Israel ;
Melo, Mariana de Carvalho ;
do Monte-Neto, Rubens Lima ;
da Silva-Pereira, Rosiane A. ;
Alves, Pedro Augusto ;
Fonseca, Cristina Toscano .
JOURNAL OF INFECTION AND PUBLIC HEALTH, 2023, 16 (07) :1081-1088
[19]   A fully integrated duplex RT-LAMP device for the detection of viral infections [J].
Mytzka, Nicolas ;
Arbaciauskaite, Skaiste ;
Sandetskaya, Natalia ;
Mattern, Kai ;
Kuhlmeier, Dirk .
BIOMEDICAL MICRODEVICES, 2023, 25 (04)
[20]   Optimization and Benchmarking of RT-LAMP-CRISPR-Cas12a for the Detection of SARS-CoV-2 in Saliva [J].
Lynch, Courtney R. H. ;
Drummond, Revel S. M. ;
Jelley, Lauren ;
Baker, Lauren ;
Smit, Erasmus ;
Fleming, Rachel ;
Billington, Craig .
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, 2025, 26 (05)
12345