Multiplex competitive annealing mediated isothermal amplification with high fidelity DNA polymerase (HiFi-CAMP).

Autor: Zhao Y; Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, China., Wan Z; Medical Laboratory of Taizhou Fourth People's Hospital, Taizhou, 225300, China., Zhang M; Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, China., Li B; Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, China., Zhang X; Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, China., Tian W; Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, China., Li YY; Department of Dermatology and Venereology, First Affiliated Hospital of Kunming Medical University, Kunming, 650032, China., Zhang C; Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, China. Electronic address: chiyu_zhang1999@163.com.
Jazyk: angličtina
Zdroj: Talanta [Talanta] 2024 Dec 01; Vol. 280, pp. 126698. Date of Electronic Publication: 2024 Aug 12.
DOI: 10.1016/j.talanta.2024.126698
Abstrakt: Various isothermal amplification methods have been developed for point-of-care testing (POCT) of various infectious diseases. Here, we proposed a novel isothermal amplification method, named as 5'-half complementary primers mediated isothermal amplification (HCPA). Because of the similarity of our method to the previous method competitive annealing mediated isothermal amplification (CAMP) in primer design, we also use the name CAMP for our method. We demonstrated that CAMP is mediated by both a linear isothermal amplification pattern and a loop-mediated isothermal amplification pattern. To improve the specificity and enable multiplex detection, we further developed HiFi-CAMP method that uses a small amount of high-fidelity DNA polymerase to cut HFman probe to release fluorescent signal. The HiFi-CAMP method was demonstrated to have a good specificity and sensitivity, and fast amplification speed in detection of three human respiratory viruses, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respiratory syncytial virus A (RSV-A) and influenza A viruses (IAV). When compared with gold standard RT-qPCR assays, the HiFi-CAMP assays showed sensitivities of 90.0 %, 71.4 % and 78.1 %, specificities of 100 %, 100 % and 95.5 %, and consistencies of 93.0 %, 93.3 % and 88.2 % for SARS-CoV-2, RSV-A and IAV, respectively. Furthermore, a duplex HiFi-CAMP assay was also developed to simultaneously detect RSV-A and SARS-CoV-2. The HiFi-CAMP will provide a promising candidate for POCT diagnosis in resource-limited settings.
Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2024 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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