Abstract
Zika virus (ZIKV) is a globally emerging mosquito-transmitted flavivirus that can cause severe fetal abnormalities, including microcephaly. As such, highly sensitive, specific, and cost-effective diagnostic methods are urgently needed. Here, we report a novel electrogenerated chemiluminescence (ECL)-based immunoassay for ultrasensitive and specific detection of ZIKV in human biological fluids. We loaded polystyrene beads (PSB) with a large number of ECL labels and conjugated them with anti-ZIKV monoclonal antibodies to generate anti-ZIKV-PSBs. These anti-ZIKV-PSBs efficiently captured ZIKV in solution forming ZIKV-anti-ZIKV-PSB complexes, which were subjected to measurement of ECL intensity after further magnetic beads separation. Our results show that the anti-ZIKV-PSBs can capture as little as 1 PFU of ZIKV in 100 μl of saline, human plasma, or human urine. This platform has the potential for development as a cost-effective, rapid and ultrasensitive assay for the detection of ZIKV and possibly other viruses in clinical diagnosis, epidemiologic and vector surveillance, and laboratory research.
Highlights
Electrogenerated chemiluminescence (ECL) is a phenomenon of light emission by an excited state of species generated at electrodes that undergo high-energetic electron-transfer reactions[29]
Maximum ECL intensities appeared at 10 mM benzyl peroxide (BPO), which likely is explained by the two opposing roles of BPO has during the ECL generation
Smaller ECL intensities were observed across the entire concentration range of BPO when DBAE and TPrA were used as coreactants
Summary
Consistent with the results obtained in PBS, our ECL-based assay detected as few as 1 PFU of ZIKV in 100 μ l of human urine (Fig. 5A) and plasma (Fig. 5B) These results were complemented by measuring the ECL intensity of virus-free PSBs that remain in solution after magnetic separations (Fig. 5C,D), which show a progressive decrease of ECL-intensity with increasing amount of ZIKV present in samples. The area under the ECL curves were plotted against amount of PFUs added to the samples (Fig. 5E,F) and produced a linear curve, suggesting that the ECL-intensity of the PSB-virus-MB aggregates formed is proportional to the amount of ZIKV in the sample These results establish that the ECL-based immunoassay can detect ZIKV in clinical samples in an ultrasensitive and specific manner
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