Graduate School of Biotechnology, Kyung Hee University, Yongin, 17104, Korea.
Department of Genetics and Biotechnology, Kyung Hee University, Yongin, 17104, Korea.
Virol J. 2023 Sep 7;20(1):206. doi: 10.1186/s12985-023-02173-1.
Severe fever with thrombocytopenia syndrome (SFTS) is an infectious disease caused by the Dabie bandavirus, [or SFTS virus (SFTSV)] that has become increasingly widespread since it was first reported in 2009. The SFTSV comprises three essential single-stranded RNA gene segments, with the S segment encoding the nucleocapsid (N) protein. Since the N protein is the most abundant and stable viral protein, it is a useful diagnostic marker of infection. Various SFTSV N-protein-based detection methods have been developed. However, given the limited research on antibodies of an SFTSV N-protein, here we report the characterization of the antibodies against SFTSV N protein especially their mapping results which is essential for more efficient and optimized detection of SFTSV.
To generate SFTSV-N-protein-specific monoclonal antibodies, recombinant full-length SFTSV N protein was expressed in E. coli, and the purified N protein was immunized to mice. The binding epitope positions of the antibodies generated were identified through binding-domain mapping. An antibody pair test using a lateral flow immunoassay (LFIA) was performed to identify effective diagnostic combinations of paired antibodies.
Nine monoclonal antibodies specific for the SFTSV N protein were generated. Antibodies #3(B4E2) and #5(B4D9) were specific for sequential epitopes, while the remainder were specific for conformational epitopes. Antibody #4(C2G1) showed the highest affinity for the SFTSV N protein. The binding domain mapping results indicated the binding regions of the antibodies were divided into three groups. The antibody pair test demonstrated that #3(B4E2)/#4(C2G1) and #4(C2G1)/#5(B4D9) were effective antibody pairs for SFTSV diagnosis.
Effective virus detection requires at least two strong antibodies recognizing separate epitope binding sites of the virus antigen. Here, we generated SFTSV-N-protein-specific monoclonal antibodies and subsequently performed epitope mapping and an antibody pair test to enhance the diagnostic efficiency and accuracy of SFTSV. Confirmation of epitope mappings and their combination immune response to the N protein provide valuable information for effective detection of SFTSV as well as can respond actively to detect a variant SFTSV.
严重发热伴血小板减少综合征（SFTS）是一种传染病，由大别山病毒（或 SFTS 病毒（SFTSV））引起，自 2009 年首次报告以来，其传播范围越来越广。SFTSV 由三个必需的单链 RNA 基因片段组成，S 片段编码核衣壳（N）蛋白。由于 N 蛋白是最丰富和最稳定的病毒蛋白，因此它是感染的有用诊断标志物。已经开发了各种基于 SFTSV N 蛋白的检测方法。然而，鉴于对 SFTSV N 蛋白抗体的研究有限，我们在这里报告了针对 SFTSV N 蛋白的抗体的特征，特别是它们的映射结果，这对于更有效地检测 SFTSV 至关重要。
为了生成 SFTSV-N 蛋白特异性单克隆抗体，在大肠杆菌中表达重组全长 SFTSV N 蛋白，并使用纯化的 N 蛋白免疫小鼠。通过结合域映射确定抗体的结合表位位置。通过侧向流动免疫分析（LFIA）进行抗体对测试，以鉴定有效的配对抗体诊断组合。
生成了针对 SFTSV N 蛋白的 9 种单克隆抗体。抗体 #3（B4E2）和 #5（B4D9）针对连续表位，而其余的则针对构象表位。抗体 #4（C2G1）对 SFTSV N 蛋白的亲和力最高。结合域映射结果表明，抗体的结合区域分为三组。抗体对测试表明，#3（B4E2）/#4（C2G1）和 #4（C2G1）/#5（B4D9）是 SFTSV 诊断的有效抗体对。
有效的病毒检测至少需要两个强抗体，以识别病毒抗原的不同表位结合位点。在这里，我们生成了 SFTSV-N 蛋白特异性单克隆抗体，随后进行了表位映射和抗体对测试，以提高 SFTSV 的诊断效率和准确性。确认表位图谱及其对 N 蛋白的组合免疫反应为有效检测 SFTSV 提供了有价值的信息，并且可以积极响应以检测变体 SFTSV。
