Development of two antigen-binding fragments to a conserved linear epitope of human adenovirus and their application in immunofluorescence

Zhenwei Liu,Huaying Chen,Yuting Xian,Rong Zhou,Xingui Tian,Wenkuan Liu,Weilue Chen,Ellen R Goldman

doi:10.1371/journal.pone.0219091

Abstract

Detection of human adenoviruses (HAdVs) in nasopharyngeal swab samples by immunofluorescence assay (IFA) will be valuable for diagnosing HAdV infection, which is a leading cause of severe respiratory tract disease, and will help in curbing the spread of HAdV. Monoclonal antibodies employed in IFA for HAdV detection should ideally target highly conserved epitope types. Here, we describe the development of two antigen-binding fragments (Fabs) with specific reactivity to HAdV using phage antibody library technology. When tested with IFA, both Fabs recognized cells infected with several types of HAdV, some of which have been identified in epidemics globally, or associated with outbreaks of severe or fatal acute respiratory diseases. The specificity and cross-reactivity of both Fabs to HAdVs indicated that the generated Fabs could be applied in the development of IFAs to detect HAdVs. Both Fabs bound to the knob proteins, as shown by chemiluminescence enzyme immunoassay and western blot. In addition, epitope mapping showed that both Fabs recognized a conserved linear epitope among several types of HAdV. Two different Fabs recognized the same epitope, suggesting that the epitope triggered the production of at least two kinds of antibodies in the body. The generated Fabs exerted no neutralization against HAdVs. The results demonstrate that both Fabs bind to an epitope that plays no role in neutralization of HAdV.

Highlights

Human adenovirus (HAdV) is a non-enveloped virus with an icosahedral shaped capsid that consists of three major proteins and four minor proteins[1]
Since there are many types of HAdV, Monoclonal antibodies (mAbs) employed in diagnostic reagents for HAdV detection should ideally target highly conserved epitope types in order to minimize the risk of failure in detecting new emergent types, as recombination is an accepted feature of HAdV evolution
We investigated the cross-reactivity of both Fabs to different types of HAdV using chemiluminescence enzyme immunoassay (CLEIA) and immunofluorescence assay (IFA), and the results demonstrated that both Fabs strongly reacted with HAdV-B3, B7, B11, B14, B55, and E4

Summary

Introduction

Human adenovirus (HAdV) is a non-enveloped virus with an icosahedral shaped capsid that consists of three major proteins (fiber, hexon, and penton base proteins) and four minor proteins[1]. The fiber, hexon, and penton base proteins of HAdV have been shown to be the antigens that cause the body to produce antibodies[2]. The fiber protein is a trimeric complex composed of three domains: an N-terminus tail, a rod-like shaft, and a globular knob at the C-. Fabs to a conserved linear epitope of human adenovirus design, data collection and analysis, decision to publish, or preparation of the manuscript

Methods

Results

Conclusion