Epitope Sequences in Dengue Virus NS1 Protein Identified by Monoclonal Antibodies.

Leticia Rocha,Viviane Botosso,Alexander Precioso,Ricardo Palacios,Thais Mitsunari,Daniela Luz,Lennon Pereira,Bruna Caetano,Neuza Gallina,Juliana Polatto,Rubens Alves,Luís Ferreira,Vanessa Silveira,Danielle Oliveira,Roxane Piazza,Celso Granato,Jaime Amorim

doi:10.3390/antib6040014

Abstract

Dengue nonstructural protein 1 (NS1) is a multi-functional glycoprotein with essential functions both in viral replication and modulation of host innate immune responses. NS1 has been established as a good surrogate marker for infection. In the present study, we generated four anti-NS1 monoclonal antibodies against recombinant NS1 protein from dengue virus serotype 2 (DENV2), which were used to map three NS1 epitopes. The sequence 193AVHADMGYWIESALNDT209 was recognized by monoclonal antibodies 2H5 and 4H1BC, which also cross-reacted with Zika virus (ZIKV) protein. On the other hand, the sequence 25VHTWTEQYKFQPES38 was recognized by mAb 4F6 that did not cross react with ZIKV. Lastly, a previously unidentified DENV2 NS1-specific epitope, represented by the sequence 127ELHNQTFLIDGPETAEC143, is described in the present study after reaction with mAb 4H2, which also did not cross react with ZIKV. The selection and characterization of the epitope, specificity of anti-NS1 mAbs, may contribute to the development of diagnostic tools able to differentiate DENV and ZIKV infections.

Highlights

Dengue fever is an important mosquito-borne and the most prevalent and costly arbovirus affecting humans, caused by one of the four serotypes of dengue virus (DENV 1–4) [1]
Fusion of popliteal lymph node cells, from mice immunized with Dengue virus serotype 2 (DENV2) rNS1, with a non-Ig-secreting or synthesizing line derived from a cell line created by fusing a BALB/c mouse spleen cell and the mouse myeloma P3X63Ag8 (SP2/O-Ag14) mouse myeloma cells, generated 25 secretory hybridomas
Four hybridomas were selected by enzyme-linked immunosorbent assay (ELISA) and sub cloned by limiting dilution and named as 4F6, 4H2, 4H1BC, and 2H5

Summary

Introduction

Dengue fever is an important mosquito-borne and the most prevalent and costly arbovirus affecting humans, caused by one of the four serotypes of dengue virus (DENV 1–4) [1]. A large number of dengue epidemics have occurred, which resulted in enormous economic and human loss in parts of Asia and South America [2,3]. NS1 is the first nonstructural protein to be translated and is essential to virus replication [5]. It is a conserved N-linked glycoprotein with a variable molecular mass of 46–55 kDa, which depends on its glycosylation status [6]. The NS1 protein can be found as a dimer associated with vesicular compartments within the cell, where it plays an important role as an essential cofactor in the virus replication process [7]. NS1 can be secreted into the extracellular space as a hexameric lipoprotein particle [8] that interacts with several plasma proteins [9,10]

Methods

Results

Conclusion