In this study, we developed a high-throughput and multiplex IgG microsphere immunoassay (MIA) using the NS1 proteins of DENV1-DENV4, ZIKV and West Nile virus (WNV) to test samples from reverse-transcription-polymerase-chain reaction-confirmed cases, including primary DENV1, DENV2, DENV3, WNV and ZIKV infections, secondary DENV infection, and ZIKV infection with previous DENV infection.
The cellular response to the recombinant NS1 protein of West Nile virus (NS1<sup>WNV</sup>) was studied using three different cell types: Vero E6 simian epithelial cells, SH-SY5Y human neuroblastoma cells, and U-87MG human astrocytoma cells.
A usual approach in this regard is the use of antibodies to nonstructural proteins as markers of nonvaccinated, infected animals, and the nonstructural NS1 protein of WNV has been proposed as a candidate for such a marker.
A stable 293T<sub>NS1</sub> cell line expressing NS1 was selected for trans-supplying NS1 protein to support the replication of WNV-ΔNS1 virus and WNV-ΔNS1-Gluc reporter virus with large-fragment deletion of NS1.
To this end we developed a bi-parametric serological microarray using recombinant NS1 proteins from Tick-borne encephalitis virus and West Nile virus coupled to a low-cost, label-free detection device based on the Reflective Phantom Interface (RPI) principle.
Plasmids to express premembrane/envelope (prM/E) or NS1 proteins of six medically important flaviviruses, including dengue viruses (DENV-1 to DENV-4), West Nile virus (WNV), and Japanese encephalitis virus (JEV), were constructed.
We generated an NS1-specific monoclonal antibody that cross-reacts with the West Nile virus (WNV) NS1 protein by immunizing mice with recombinant JEV NS1.
By screening and identifying the specific WNVNS1 protein-derived peptides recognized by serum antibodies elicited by immunization with WNVNS1 protein and by JEV challenge, we found after JEV challenge will induce several new epitopes, but which epitope primarily contribute to antibody-mediated cross protection need further evaluation.
We described previously mAbs against the non-structural NS1 protein of the West Nile virus (WNV) that were protective in mice when administered pre- or post-infection of WNV.
The search revealed a stretch of 12 amino acids in the NS1 protein of West Nile virus with a high degree of homology to the GAD65 region (an isoform of GAD) containing the PEVKEK motif.