Sidney J, Oseroff C, Southwood S, Wall M, Ishioka G, Koning F, Sette A
Cytel, San Diego, CA 92121.
J Immunol. 1992 Oct 15;149(8):2634-40.
The DR3-restricted peptide, MT 65 kDa 3-13, was used to develop a DR3-specific binding assay. The binding activity detected was strictly pH-dependent, in that it was optimal in the pH 4 to 5 range, and no activity was detected at neutral pH. By means of affinity chromatography purifications and the use of DR3-transfected fibroblasts, it was shown that no cross-reactivity exists at the level of DR52a molecules, thus allowing use of only partially purified DR3/DR52a mixtures in high throughput binding assays. The immunologic relevance of the assay established was also verified by examining the correlation between DR3 restriction and binding for a panel of DR-restricted peptides. When the structural requirements for peptide DR3 interactions were further examined by using panels of analogues of two different epitopes (Myo 132-151 and TT 830-843), it was found that DR3 molecules recognized a peptide motif distinct from the one recognized by the other major DR beta 1 alleles.
受DR3限制的肽MT 65 kDa 3 - 13被用于开发一种DR3特异性结合检测方法。检测到的结合活性严格依赖于pH值，即在pH 4至5范围内最佳，在中性pH值下未检测到活性。通过亲和层析纯化以及使用转染了DR3的成纤维细胞，结果表明在DR52a分子水平不存在交叉反应，因此在高通量结合检测中仅使用部分纯化的DR3/DR52a混合物即可。通过检查一组DR限制肽的DR3限制与结合之间的相关性，也验证了所建立检测方法的免疫学相关性。当使用两种不同表位（Myo 132 - 151和TT 830 - 843）的类似物组进一步研究肽与DR3相互作用的结构要求时，发现DR3分子识别的肽基序与其他主要DRβ1等位基因识别的不同。
