抄録
BACKGROUND: Zika virus (ZIKV) is a mosquito-borne flavivirus. ZIKV infection is considered to be the cause of microcephaly in fetuses and newborns and Guillain-Barre syndrome by recent epidemics of ZIKV infection in Brazil. The diagnostic of ZIKV infection is performed in laboratories by nucleic acid amplification tests (rRT-PCR) and serological tests. Since flaviviruses are highly conserved, serologic tests are difficult to use for diagnosis due to cross-reactivity to other flavivirus. The rRT-PCT is mainly used for the diagnosis but requires special techniques and equipment and takes time. Thus, simpler diagnostic tests are desired. Since flavivirus nonstructural protein 1 (NS1) is secreted from virus-infected cells into the bloodstream, this protein is known to remain detectable in the blood even after the viral genome has disappeared and is useful as an infectious disease marker. Therefore, we developed rapid immunochromatographic kits for detection of ZIKV NS1.
METHODS: Monoclonal antibodies (MoAbs) specific to ZIKV NS1 were established by immunizing mice with recombinants or synthetic peptides. Characterizations of the MoAbs were investigated by ELISA using recombinants. Rapid immunochromatographic kits were developed using established MoAbs and then evaluated using recombinants, culture supernatants of ZIKV-infected cells and clinical specimens.
RESULTS: We established three MoAbs (2-42, 1-78 and 84F9) with high reactivity and specificity to ZIKV NS1. Rapid immunochromatographic kits were developed with available three MoAbs. These rapid kits could detect NS1 derived from two ZIKV strains (African and Asian strain) using both recombinants and culture supernatants of ZIKV-infected cell lines. Detection sensitivity of our kits was 125 pg/mL using recombinant NS1. In the testing against five clinical specimens collected in Brazil in 2016, which were confirmed ZIKV infection by rRT-PCR (Ct=19.4-21.6), our kits could detect all specimens as positive and also detect NS1 in over 2000-fold diluted sample within 15 min. Further, the kits showed no cross-reactivity against specimens infected with dengue virus 1 to 4 containing NS1 at high level.
CONCLUSIONS: Our kits constructed in this study could detect NS1 in ZIKV-infected specimens at high sensitivity and specificity and might be useful for clinical application to acute-phase of ZIKV infection.
