Optimization of loop-mediated isothermal amplification (LAMP) assay for robust visualization in SARS-CoV-2 and emerging variants diagnosis

Zhen Luo; Chunhong Ye; Heng Xiao; Jialing Yin; Yicong Liang; Zhihui Ruan; Danju Luo; Daolong Gao; Qiuping Tan; Yongkui Li; Qiwei Zhang; Weiyong Liu; Jianguo Wu

doi:10.1016/j.ces.2022.117430

Optimization of loop-mediated isothermal amplification (LAMP) assay for robust visualization in SARS-CoV-2 and emerging variants diagnosis

Chem Eng Sci. 2022 Apr 6:251:117430. doi: 10.1016/j.ces.2022.117430. Epub 2022 Jan 13.

Authors

Zhen Luo^{1

2}, Chunhong Ye¹, Heng Xiao^{1

2}, Jialing Yin¹, Yicong Liang¹, Zhihui Ruan², Danju Luo³, Daolong Gao⁴, Qiuping Tan⁴, Yongkui Li^{1

2}, Qiwei Zhang^{1

2}, Weiyong Liu⁵, Jianguo Wu^{1

2

4}

Affiliations

¹ Guangdong Provincial Key Laboratory of Virology, Institute of Medical Microbiology, Jinan University, Guangzhou 510632, China.
² Foshan Institute of Medical Microbiology, Foshan 528315, China.
³ Department of Pathology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
⁴ Guangdong Longfan Biological Science and Technology Company, Shunde District, Foshan 528315, China.
⁵ Tongji Hospital of Huazhong University of Science and Technology, Wuhan 430030, China.

Abstract

Loop-mediated isothermal amplification (LAMP) is widely used in detection of pathogenic microorganisms including SARS-CoV-2. However, the performance of LAMP assay needs further exploration in the emerging SARS-CoV-2 variants test. Here, we design serials of primers and select an optimal set for LAMP-based on SARS-CoV-2 N gene for a robust and visual assay in SARS-CoV-2 diagnosis. The limit of detectable template reaches 10 copies of N gene per 25 μL reaction at isothermal 58℃ within 40 min. Importantly, the primers for LAMP assay locate at 12 to 213 nt of N gene, a highly conservative region, which serves as a compatible test in emerging SARS-CoV-2 variants. Comparison to a commercial qPCR assay, this LAMP assay exerts the high viability in diagnosis of 41 clinical samples. Our study optimizes an advantageous LAMP assay for colorimetric detection of SARS-CoV-2 and emerging variants, which is hopeful to be a promising test in COVID-19 surveillance.

Keywords: COVID-19, coronavirus disease 2019; CRISPR, clustered regularly interspaced short palindromic repeats; Coronavirus disease 2019 (COVID-19) pandemic; Ct, threshold cycle; Emerging SARS-CoV-2 variants; IVD, in-vitro diagnosis; LAMP, Loop-mediated isothermal amplification; Loop-mediated isothermal amplification (LAMP); NGS, next-generation sequencing; POC, point-of-care; RT-qPCR, real-time reverse transcriptase quantitative polymerase chain reaction; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnosis; VOC, variants of concern.