Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization.
Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization.
复制标题
DOI：
10.1002/0471142735.im0510s106
复制
发表时间：
2014-08-01
影响因子：
--
通讯作者：
Stern, Lawrence J
中科院分区：
文献类型：
--
作者：
Yin, Liusong;Stern, Lawrence J
肽与主要组织相容性复合物II类（MHCII）分子的结合是抗原呈递和CD4+ T细胞表位选择的关键过程。本单元描述了一种相当简单但功能强大的基于荧光偏振的结合竞争测定法，用于测量肽与可溶性重组MHCII分子的结合。目标肽与MHCII分子的结合基于其抑制荧光标记的探针肽结合的能力来评估，结合强度表征为IC 50（50%抑制探针肽结合所需的浓度）。讨论了与该方法相关的数据分析。此外，该单元还包括使用胺反应性探针进行荧光标记肽的支持方案。该方案的优点是它允许简单、快速和高通量地测量大量肽与MHCII分子的结合。
Peptide binding to major histocompatibility complex class II (MHCII) molecules is a key process in antigen presentation and CD4+ T cell epitope selection. This unit describes a fairly simple but powerful fluorescence polarization-based binding competition assay to measure peptide binding to soluble recombinant MHCII molecules. The binding of a peptide of interest to MHCII molecules is assessed based on its ability to inhibit the binding of a fluorescence-labeled probe peptide, with the strength of binding characterized as IC50 (concentration required for 50% inhibition of probe peptide binding). Data analysis related to this method is discussed. In addition, this unit includes a support protocol for fluorescence labeling peptide using an amine-reactive probe. The advantage of this protocol is that it allows simple, fast, and high throughput measurements of binding for a large set of peptides to MHCII molecules.