Multiplexed Magnetofluorescent Bioplatform for the Sensitive Detection of SARS-CoV-2 Viral RNA without Nucleic Acid Amplification.
Riham ZayaniDorra RezigWasfi FaresMouna MarrakchiMakram EssafiNoureddine RaouafiPublished in: Analytical chemistry (2021)
Rapid and sensitive detection of SARS-CoV-2 virus genetic material is of paramount importance to mitigate the COVID-19 pandemic outbreak and lower the death toll. Herein, we report the design of a magnetofluorescent bioplatform for the direct and specific detection of the viral RNA of SARS-CoV-2 in the total RNA extracted from nasopharyngeal swabs of COVID-19-positive patients. A higher fluorescence response was achieved using two capture probes tethered to magnetic beads using a biotin/streptavidin linkage, targeting two specific sites in the ORF1a and S genes. Two horseradish peroxidase (HRP)-conjugated reporter sequences, complementary to the loci of the S and N genes, were used to reveal the presence of the viral RNA through the oxidation of o-phenylenediamine to fluorescent 2,3-diaminophenazine. Under optimal conditions, the bioplatform showed high selectivity and sensitivity and was able to detect as low as 0.01 ng of viral RNA (1 × 103 copies/μL) with a linear dynamic range varying from 0.01 to 3.0 ng (1 × 103 to 9 × 107 copies/μL). The bioplatform was also able to discriminate the SARS-CoV-2 RNA from those of other related viruses such as hepatitis C, West Nile, measles, and non-polio viruses. Furthermore, the developed biosensor was validated in 46 clinical samples (36 COVID-19-positive patients and 10 COVID-19-negative subjects, as assessed with the gold standard RT-qPCR method). Both sensitivity and specificity of the developed method reached 100%. Finally, making such a simple and specific method available in the field, at a primary point of care, can better help the detection of SARS-CoV-2 infection in low-resource settings.
Keyphrases
- sars cov
- nucleic acid
- sensitive detection
- respiratory syndrome coronavirus
- loop mediated isothermal amplification
- genome wide
- quantum dots
- end stage renal disease
- ejection fraction
- chronic kidney disease
- newly diagnosed
- prognostic factors
- gold nanoparticles
- dna methylation
- hydrogen peroxide
- gene expression
- single molecule
- nitric oxide
- crispr cas
- copy number
- patient reported outcomes
- human immunodeficiency virus
- cancer therapy
- liquid chromatography
- fluorescence imaging
- real time pcr
- patient reported
- electron transfer