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Evaluation of real-time NASBA assay for the detection of SARS-CoV-2 compared with real-time PCR.
Kia, Vahid; Tafti, Ali; Paryan, Mahdi; Mohammadi-Yeganeh, Samira.
  • Kia V; Department of Medical Biotechnology, School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran.
  • Tafti A; Department of Medical Biotechnology, Faculty of Medicine, Arak University of Medical Science, Arak, Iran.
  • Paryan M; Department of Research and Development, Production and Research Complex, Pasteur Institute of Iran, Tehran, Iran. mparyan@gmail.com.
  • Mohammadi-Yeganeh S; Medical Nanotechnology and Tissue Engineering Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran. smyeganeh@gmail.com.
Ir J Med Sci ; 2022 Jun 06.
Article in English | MEDLINE | ID: covidwho-2266311
ABSTRACT

PURPOSE:

In January 2020, the COVID-19 pandemic started and has severely affected all countries around the world. The clinical symptoms alone are not sufficient for a proper diagnosis. Thus, molecular tests are required. Various institutes and researchers developed real-time PCR-based methods for the detection of the virus. However, the method needs expensive equipment. In the present study, we developed a real-time NASBA assay for the detection of SARS-CoV-2.

METHODS:

Primers and molecular beacon probes for RdRp and N genes were designed. In silico analysis showed that primers and the probes were specific for SARS-CoV-2. The standard samples with known copy numbers of the virus were tested using the NASBA assay and an FDA-approved real-time PCR kit. A series of standard samples were prepared and tested. Clinical sensitivity, precision analysis, and clinical assessment of the assay were performed.

RESULTS:

The limit of detection of the assay was 200 copies/mL. The clinical sensitivity of the assay was 97.64%. The intra-assay and inter-assay for both N and RdRp genes were less than 5% and 10%, respectively. Clinical assessment of the assay showed that the positive agreement rate and negative agreement rate of the assays were determined to be 97.64% and 100%, respectively.

CONCLUSIONS:

The results of the present study show that the developed real-time NASBA is a sensitive and specific method for the detection of SARS-CoV-2 and is comparable with real-time PCR. NASBA is an isothermal signal amplification method, and if stand-alone fluorescent readers are available, the real-time NASBA can be used without the need for expensive thermocyclers. In addition compared to other isothermal methods like LAMP, the primer design is straightforward. Thus, real-time NASBA could be a suitable method for inexpensive SARS-CoV-2 detection.
Keywords

Full text: Available Collection: International databases Database: MEDLINE Type of study: Diagnostic study / Experimental Studies / Prognostic study Language: English Year: 2022 Document Type: Article Affiliation country: S11845-022-03046-2

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Full text: Available Collection: International databases Database: MEDLINE Type of study: Diagnostic study / Experimental Studies / Prognostic study Language: English Year: 2022 Document Type: Article Affiliation country: S11845-022-03046-2