Developing novel liquid biopsy by selective capture of viral RNA on magnetic beads to detect COVID-19

Document Type : Original Article

Authors

1 Medical Genetics Research Center, Mashhad University of Medical Sciences, Mashhad, Iran

2 Department of Medical Genetics, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran

3 Genetics Research Unit, Reza Radiotherapy and Oncology Center, Mashhad, Iran

4 Antimicrobial Resistance Research Center, Mashhad University of Medical Sciences, Mashhad, Iran

5 Department of Animal Science, Faculty of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran

6 Department of Medical Genetics, Shahid Sadoughi University of Medical Sciences, Yazd, Iran

7 Lung Diseases Research Center, Mashhad University of Medical Sciences, Mashhad, Iran

Abstract

Objective(s): Early, specific, and sensitive detection methods of COVID-19 are essential for force stopping its worldwide infection. Although CT images of the lung and/or viral RNA extraction followed by real-time reverse-transcriptase-polymerase chain reaction (rRT-PCR) are widely used; they have some limitations. Here, we developed a highly sensitive magnetic bead-based viral RNA extraction assay followed by rRT-PCR. 
Materials and Methods: Case group included oropharyngeal/nasopharyngeal and blood samples from 30 patients diagnosed positive by PCR test for COVID-19 and control group included 30 same samples from COVID-19 negative PCR test individuals. RNA was extracted, using viral RNA extraction kit as well as using our hand-made capture bead-based technique. A one-step cDNA synthesis and Real Time PCR was conducted. A two-step comparison of the different viral RNA extraction methods for oropharyngeal/nasopharyngeal and blood samples was performed. Student t-test was applied with a P<0.05 considered statistically significant. 
Results: In the case group, all 30 mucosal samples extracted either with viral RNA extraction kit or with beads-based assay were COVID-19 positive although in the latter category, Cqs were much lower. Although 43% of plasma samples extracted by bead-based method were found to be positive but no plasma samples extracted with column-based kit were detected positive by Real Time PCR. 
Conclusion: Bead-based RNA extraction method can reduce RNA loss by its single-tube performance and enhance the test sensitivity. It is also more sensitive to lower viral loads as shown in the detection of blood samples and the lower Cqs of mucosal samples.

Keywords

References

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Iranian Journal of Basic Medical Sciences
Volume 25, Issue 6
June 2022
Pages 762-766

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