Department of Chemical Biology , Helmholtz Centre for Infection Research and German Centre for Infection Research (DZIF) , 38124 Braunschweig , Germany.
Division of Microbiology and Animal Hygiene , Georg-August-University , 37077 Göttingen , Germany.
Anal Chem. 2019 Sep 3;91(17):11030-11037. doi: 10.1021/acs.analchem.9b01291. Epub 2019 Aug 14.
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.
猕猴疱疹病毒或 B 病毒（BV）是一种人畜共患病原体，如果传播且未得到治疗，会导致人类的高死亡率。在这里，BV 被用作案例研究，以建立一种基于合成肽的高通量血清学检测的两步法程序。在步骤 1 中，通过对 42 份猴血清（其中 30 份通过 ELISA 检测为 BV 阳性）进行肽微阵列分析，发现了针对 369 种不同肽的 1148 种反应。后者可以在 BV 的六种不同糖蛋白（gB、gC、gD、gG、gH 和 gL）的 142 个不同抗体靶区（ATR）中进行分组。新检测到的 ATR 数量之所以如此之多，部分原因是采用了一种新的预处理方案，该方案减少了血清成分与微阵列纤维素基质的非特异性结合。在步骤 2 中，合成了对应于 8 个具有特别高抗原性的 ATR 的可溶性肽，并将其偶联到荧光标记的珠上，随后将其用于免疫化学珠流分析中。它们的结果与用作参考的 ELISA 结果一致。因此，现在可以方便、快速和经济地对任意大的猕猴群体进行 BV 感染筛查。该研究表明，从用于表位发现的二维高分辨率肽阵列技术平台切换到可用于血清学应用的现成珠阵列平台是可行的。
