Abstract
Developing rapid and sensitive diagnostic methods for dengue virus (DENV) infection is of prime priority because DENV infection is the most prevalent mosquito-borne viral disease. This work proposes an electrochemical impedance spectroscopy (EIS)-based genosensor for the label-free and nucleic acid amplification-free detection of extracted DENV RNA intended for a sensitive diagnosis of DENV infection. A concentration ratio of 0.04 mM 6-mercaptohexanoic acid (MHA) to 1 mM 6-mercapto-1-hexanol (MCH) was selected to modify thin-film gold electrodes as a link to control the coverage of self-designed probe DNA (pDNA) at a density of 4.5 ± 0.4 × 1011 pDNA/cm2. The pDNA/MHA/MCH-modified genosensors are proven to improve the hybridization efficiency of a synthetic 160-mer target DNA (160mtDNA) with a 140-mer electrode side overhang as compared to other MHA/MCH ratio-modified genosensors. The MHA(0.04 mM)/MCH(1 mM)-modified genosensors also present good hybridization efficiency with the extracted DENV serotype 1 (DENV1) RNA samples, having the same electrode side overhangs with the 160mtDNA, showing a low detection limit of 20 plaque forming units (PFU)/mL, a linear range of 102–105 PFU/mL and good selectivity for DENV1. The pDNA density-controlled method has great promise to construct sensitive genosensors based on the hybridization of extracted DENV nucleic acids.
Highlights
The epidemic of dengue, malaria, chikungunya, yellow fever, West Nile virus and Japanese encephalitis, notorious diseases transmitted by mosquitoes [1], have been propelled by climate warming, globalization and increasing international activity [2,3,4]
The results indicated that the probe DNA (pDNA) density of mercaptohexanoic acid (MHA)(0.04
The ratio-varied Mercaptohexanoic acid/6-mercapto-1-hexanol (MHA/MCH) binary Self-Assembled Monolayer (SAM) can be used as a link to control the density of immobilized pDNA
Summary
The epidemic of dengue, malaria, chikungunya, yellow fever, West Nile virus and Japanese encephalitis, notorious diseases transmitted by mosquitoes [1], have been propelled by climate warming, globalization and increasing international activity [2,3,4]. Developing rapid and sensitive diagnostic methods for DENV infection from the onset of febrile symptoms is very important in dengue fever prevention and treatment. Primary infection of patients can be caused by any one of the four serotypes of DENV (DENV1, 2, 3, and 4) and secondary infection of these patients with a heterologous serotype increases the risk of progressing into severe dengue due to antibody-dependent enhancement [22,23] compared with NS1 or anti-DENV IgM, DENV particles are the first to be detected in dengue patient serum post symptom onset [21] The serotype-specific detection of DENV will be more time-effective for DENV diagnosis and more meaningful for clinical treatments
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