Abstract
Serologic tests to detect specific IgGs to antigens related to viral infections are urgently needed for diagnostics and therapeutics. We present a diagnostic method for serotype-specific IgG identification of dengue infection by a competitive enzyme-linked immunosorbent assay (ELISA), using high-affinity unnatural-base-containing DNA (UB-DNA) aptamers that recognize the four categorized serotypes. Using UB-DNA aptamers specific to each serotype of dengue NS1 proteins (DEN-NS1), we developed our aptamer–antibody sandwich ELISA for dengue diagnostics. Furthermore, IgGs highly specific to DEN-NS1 inhibited the serotype-specific NS1 detection, inspiring us to develop the competitive ELISA format for dengue serotype-specific IgG detection. Blood samples from Singaporean patients with primary or secondary dengue infections confirmed the highly specific IgG detection of this format, and the IgG production initially reflected the serotype of the past infection, rather than the recent infection. Using this dengue competitive ELISA format, cross-reactivity tests of 21 plasma samples from Singaporean Zika virus-infected patients revealed two distinct patterns: 8 lacked cross-reactivity, and 13 were positive with unique dengue serotype specificities, indicating previous dengue infection. This antigen-detection ELISA and antibody-detection competitive ELISA combination using the UB-DNA aptamers identifies both past and current viral infections and will facilitate specific medical care and vaccine development for infectious diseases.
Highlights
The recent COVID-19 (Coronavirus Disease 2019) pandemic has highlighted the importance of serologic tests, for infectious disease diagnostics complementary to PCR and biomarker-detection tests[1,2,3,4]
We serendipitously found that the unnatural-basecontaining DNA (UB-DNA) aptamer binding to dengue non-structural protein 1 (NS1) proteins (DEN-NS1) is serotype- inhibited when anti-DEN-NS1 IgG antibodies were present in patient blood samples
PZ-20 revealed another discrepancy between our competitive enzyme-linked immunosorbent assay (ELISA) and the SD BIOLINE test. These differences might result from the low sensitivity of the commercially available paper-based assay formats and the differences in the targeting dengue viral (DENV) antigens. These results suggest that our dengue competitive ELISA is not cross-reactive with anti-Zika virus (ZIKV)-NS1 IgGs and can detect prior DENV infections in patients currently infected by ZIKV
Summary
The recent COVID-19 (Coronavirus Disease 2019) pandemic has highlighted the importance of serologic tests, for infectious disease diagnostics complementary to PCR and biomarker-detection tests[1,2,3,4]. Serologic tests enable the detection of viral-specific antibodies, mainly IgM and IgG, produced in the body by responses to current and past infections. Such tests are useful for the diagnoses of infection, including surveys of disease transmission, infection spread, and acquired immunity, as well as evaluations of vaccine development. Most typical serologic tests are methods to detect antibodies by binding to viral-related antigens using lateral flow devices and the enzyme-linked immunosorbent assay (ELISA) or chemiluminescent immunoassay (CIA) format[5,6,7,8] In these methods, the sensitivity and specificity of detection due to the cross-reactivity with other related diseases are one of the key issues, as they cause false-negative and false-positive test results. The development of detection methods for current infection and past infection, including the serotype identification and quantification, is an urgent worldwide t ask[31,34]
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