Zika Virus Non-Structural Protein 1 Antigen-Capture Immunoassay.

Brandon J Beddingfield,Refugio Robles-Sikisaka,Jessica N Hartnett,Robert F Garry,Kristian G Andersen,Russell B Wilson,Peter C Kulakosky,Nathan D Grubaugh,Maria-Zunilla Nunez,Argelia Aybar,Cesar D Fermin

doi:10.3390/v13091771

Abstract

Infection with Zika virus (ZIKV), a member of the Flavivirus genus of the Flaviviridae family, typically results in mild self-limited illness, but severe neurological disease occurs in a limited subset of patients. In contrast, serious outcomes commonly occur in pregnancy that affect the developing fetus, including microcephaly and other major birth defects. The genetic similarity of ZIKV to other widespread flaviviruses, such as dengue virus (DENV), presents a challenge to the development of specific ZIKV diagnostic assays. Nonstructural protein 1 (NS1) is established for use in immunodiagnostic assays for flaviviruses. To address the cross-reactivity of ZIKV NS1 with proteins from other flaviviruses we used site-directed mutagenesis to modify putative epitopes. Goat polyclonal antibodies to variant ZIKV NS1 were affinity-purified to remove antibodies binding to the closely related NS1 protein of DENV. An antigen-capture ELISA configured with the affinity-purified polyclonal antibody showed a linear dynamic range between approximately 500 and 30 ng/mL, with a limit of detection of between 1.95 and 7.8 ng/mL. NS1 proteins from DENV, yellow fever virus, St. Louis encephalitis virus and West Nile virus showed significantly reduced reactivity in the ZIKV antigen-capture ELISA. Refinement of approaches similar to those employed here could lead to development of ZIKV-specific immunoassays suitable for use in areas where infections with related flaviviruses are common.

Highlights

Due to the sequence similarity between Nonstructural protein 1 (NS1) proteins of flaviviruses, the Zika virus (ZIKV) NS1 is expected to be structurally similar to the NS1 proteins of West Nile virus (WNV), dengue virus (DENV) and other members of the flavivirus genus of the Flaviviridae
A diagnostic based on capture of WNV NS1 was developed using monoclonal antibodies (mAbs), where capture of recombinant NS1 was deemed sensitive, with a detection limit of 0.5 ng/mL soluble NS1
An antigen-capture Enzyme Linked Immunosorbent Assay (ELISA) configured with an affinity-purified polyclonal antibodies (pAbs) to ZIKV NS1 demonstrated limited cross-reactivity to NS1 of commonly circulating flaviviruses

Summary

Introduction

Zika virus (ZIKV) is named after the Zika forest in Uganda, where it was first isolated from a sentinel monkey in 1947 [1], and shortly after from mosquitoes in the same area [2]. Follow-up serological studies revealed Zika to be widespread in Africa and Asia [1,3,4]. The first reported natural ZIKV infection in humans was reported in 1964, when a scientist was infected while isolating virus from mosquitos in Uganda [4,5]. The first known incidence of Zika outside Africa and Asia was in 2007 in Yap State, Micronesia [6], in what became the first large outbreak recorded [7]. In 2013, cases of Zika began being reported in French Polynesia

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