Ph.D. Program in Medical Biotechnology, College of Medical Science and Technology, Taipei Medical University, Taipei 110301, Taiwan.
School of Medical Laboratory Science and Biotechnology, College of Medical Science and Technology, Taipei Medical University, Taipei 110301, Taiwan.
Int J Mol Sci. 2021 Apr 16;22(8):4146. doi: 10.3390/ijms22084146.
A16 (CA16) is one of the major causative agents of hand, foot, and mouth disease (HFMD). Children aged <5 years are the most affected by CA16 HFMD globally. Although clinical symptoms of CA16 infections are usually mild, severe complications, such as aseptic meningitis or even death, have been recorded. Currently, no vaccine or antiviral therapy for CA16 infection exists. Single-chain variable fragment (scFv) antibodies significantly inhibit viral infection and could be a potential treatment for controlling the infection. In this study, scFv phage display libraries were constructed from splenocytes of a laying hen immunized with CA16-infected lysate. The pComb3X vector containing the scFv genes was introduced into ER2738 and rescued by helper phages to express scFv molecules. After screening with five cycles of bio-panning, an effective scFv antibody showing favorable binding activity to proteins in CA16-infected lysate on ELISA plates was selected. Importantly, the selected scFv clone showed a neutralizing capability against the CA16 virus and cross-reacted with viral proteins in EV71-infected lysate. Intriguingly, polyclonal IgY antibody not only showed binding specificity against proteins in CA16-infected lysate but also showed significant neutralization activities. Nevertheless, IgY-binding protein did not cross-react with proteins in EV71-infected lysate. These results suggest that the IgY- and scFv-binding protein antibodies provide protection against CA16 viral infection in in vitro assays and may be potential candidates for treating CA16 infection in vulnerable young children.
CA16（CA16）是手足口病（HFMD）的主要病原体之一。全球范围内，<5 岁的儿童受 CA16 HFMD 的影响最大。尽管 CA16 感染的临床症状通常较轻，但已记录到严重的并发症，如无菌性脑膜炎甚至死亡。目前，尚无针对 CA16 感染的疫苗或抗病毒疗法。单链可变片段（scFv）抗体可显著抑制病毒感染，可能成为控制感染的潜在治疗方法。在这项研究中，从用 CA16 感染的裂解物免疫的产蛋鸡的脾细胞中构建了 scFv 噬菌体展示文库。将含有 scFv 基因的 pComb3X 载体引入 ER2738 中，并通过辅助噬菌体拯救以表达 scFv 分子。经过五轮生物淘选筛选后，选择了一种有效的 scFv 抗体，该抗体在 ELISA 板上对 CA16 感染的裂解物中的蛋白质表现出良好的结合活性。重要的是，所选的 scFv 克隆显示出对 CA16 病毒的中和能力，并与 EV71 感染的裂解物中的病毒蛋白发生交叉反应。有趣的是，多克隆 IgY 抗体不仅针对 CA16 感染的裂解物中的蛋白质表现出结合特异性，而且还显示出显著的中和活性。然而，IgY 结合蛋白与 EV71 感染的裂解物中的蛋白质没有发生交叉反应。这些结果表明，IgY 和 scFv 结合蛋白抗体在体外试验中提供了针对 CA16 病毒感染的保护作用，并且可能是治疗易受感染的幼儿 CA16 感染的潜在候选物。
