Center for Technological Development in Health (CDTS)/National Institute of Science and Technology for Innovation in Neglected Diseases Populations (INCT-IDPN), Oswaldo Cruz Foundation (FIOCRUZ), Rio de Janeiro 21040-900, RJ, Brazil.
Epidemiology and Molecular Systematics Laboratory (LEMS), Oswaldo Cruz Institute, Oswaldo Cruz Foundation (FIOCRUZ), Rio de Janeiro 21040-900, RJ, Brazil.
Int J Mol Sci. 2022 Dec 28;24(1):531. doi: 10.3390/ijms24010531.
Oral immunization with the choleric toxin (CT) elicits a high level of protection against its enterotoxin activities and can control cholera in endemic settings. However, the complete B-cell epitope map of the CT that is responsible for protection remains to be clarified. A library of one-hundred, twenty-two 15-mer peptides covering the entire sequence of the three chains of the CT protein (CTP) was prepared by SPOT synthesis. The immunoreactivity of membrane-bound peptides with sera from mice vaccinated with an oral inactivated vaccine (Schankol™) allowed the mapping of continuous B-cell epitopes, topological studies, multi-antigen peptide (MAP) synthesis, and Enzyme-Linked Immunosorbent Assay (ELISA) development. Eighteen IgG epitopes were identified; eight in the CTA, three in the CTB, and seven in the protein P. Three specific epitopes, Vc/TxA-3, Vc/TxB-11, and Vc/TxP-16, were synthesized as MAP4 and used to coat ELISA plates in order to screen immunized mouse sera. Sensitivities and specificities of 100% were obtained with the MAP4s of Vc/TxA-3 and Vc/TxB-11. The results revealed a set of peptides whose immunoreactivity reflects the immune response to vaccination. The array of peptide data can be applied to develop improved serological tests in order to detect cholera toxin exposure, as well as next generation vaccines to induce more specific antibodies against the cholera toxin.
口服免疫接种霍乱肠毒素（CT）可引发针对其肠毒素活性的高水平保护作用，并可控制地方性流行环境中的霍乱。然而，负责保护作用的 CT 的完整 B 细胞表位图谱仍有待阐明。通过 SPOT 合成制备了一个由 122 个 15 肽组成的文库，覆盖 CT 蛋白（CTP）的三条链的整个序列。用口服灭活疫苗（Schankol™）接种的小鼠血清中膜结合肽的免疫反应性允许连续 B 细胞表位的作图、拓扑研究、多抗原肽（MAP）合成和酶联免疫吸附测定（ELISA）的开发。鉴定了 18 个 IgG 表位；8 个在 CTA 中，3 个在 CTB 中，7 个在蛋白 P 中。三个特异性表位 Vc/TxA-3、Vc/TxB-11 和 Vc/TxP-16 作为 MAP4 合成，并用于包被 ELISA 板，以筛选免疫小鼠血清。用 Vc/TxA-3 和 Vc/TxB-11 的 MAP4 获得了 100%的灵敏度和特异性。结果揭示了一组肽，其免疫反应性反映了对疫苗接种的免疫反应。肽数据的数组可用于开发改进的血清学检测方法，以检测霍乱毒素暴露，以及诱导针对霍乱毒素的更特异性抗体的下一代疫苗。
