Berndt Thalén Niklas, Karlander Maximilian, Malm Magdalena, Rockberg Johan
Department of Protein Science, KTH-Royal Institute of Technology, Stockholm, Sweden.
Methods Mol Biol. 2025;2937:133-146. doi: 10.1007/978-1-0716-4591-8_8.
Precise epitope determination of therapeutic antibodies is of great value as it allows for further comprehension of the mechanism of action, therapeutic responsiveness prediction, avoidance of unwanted cross-reactivity, and vaccine design. Alanine scanning mutagenesis can provide an amino acid level resolution of a precise epitope for conformational epitopes. If the mutated antigen is presented on the surface of cells, high throughput screening is enabled without the need for protein expression and purification. Here we present a combinatorial method for rapid mapping of discontinuous epitopes by mammalian antigen display, facilitated by automated workflows for high throughput cloning and tailored software for antigen analysis and oligonucleotide design.
治疗性抗体精确表位的确定具有重要价值，因为它有助于进一步理解作用机制、预测治疗反应性、避免不必要的交叉反应以及进行疫苗设计。丙氨酸扫描诱变可为构象表位提供精确表位的氨基酸水平分辨率。如果突变抗原呈现在细胞表面，则无需蛋白质表达和纯化即可进行高通量筛选。在此，我们介绍一种通过哺乳动物抗原展示快速绘制不连续表位的组合方法，该方法借助高通量克隆的自动化工作流程以及用于抗原分析和寡核苷酸设计的定制软件得以实现。
