Serological Analysis of Herpes B Virus at Individual Epitope Resolution: From Two-Dimensional Peptide Arrays to Multiplex Bead Flow Assays

Abstract

Macacine herpesvirus or B Virus (BV) is a zoonotic agent that leads to high mortality rates in humans if transmitted and untreated. Here, BV is used as a test case to establish a two-step procedure for developing high throughput serological assays based on synthetic peptides. In step 1, peptide microarray analysis of 42 monkey sera (30 of them tested BV positive by ELISA) revealed 1148 responses against 369 different peptides. The latter could be grouped into 142 different antibody target regions (ATRs) in six different glycoproteins (gB, gC, gD, gG, gH, and gL) of BV. The high number of newly detected ATRs was made possible inter alia by a new preanalytical protocol that reduced unspecific binding of serum components to the cellulose-based matrix of the microarray. In step 2, soluble peptides corresponding to eight ATRs of particularly high antigenicity were synthesized and coupled to fluorescently labeled beads, which were subsequently employed in immunochemical bead flow assays. Their outcome mirrored the ELISA results used as reference. Hence, convenient, fast, and economical screening of arbitrarily large macaque colonies for BV infection is now possible. The study demonstrates that a technology platform switch from two-dimensional high-resolution peptide arrays used for epitope discovery to a readily available bead array platform for serology applications is feasible.