Finnern R, Pedrollo E, Fisch I, Wieslander J, Marks J D, Lockwood C M, Ouwehand W H
Department of Transfusion Medicine, University of Cambridge, UK.
Clin Exp Immunol. 1997 Feb;107(2):269-81. doi: 10.1111/j.1365-2249.1997.254-ce1127.x.
This is the first study describing recombinant human antibody fragments directed to the autoantigen proteinase 3 (PR3) from an immune B cell source. Detection of these autoantibodies has proven valid for the diagnosis and monitoring of Wegener's granulomatosis. The described antibody fragment (scFv) was isolated from a phage display library prepared from the IgG-positive splenic lymphocytes of a patient with systemic autoimmunity. The cloning strategy was designed to maintain the diversity of the antibody variable gene repertoire, and sequencing of several variable genes demonstrated that all major heavy and light chain families were represented. We found an over-representation of particular heavy chain variable domains in splenic lymphocytes which differ from the ones frequently found in peripheral blood lymphocytes. It was possible to obtain specific scFv to PR3 after a single round of selection and the binding could be inhibited by the patients' sera. Although the antibody fragments in the splenic repertoire were found to be highly mutated, it was interesting to find that the selected scFv showed only limited somatic mutation. Furthermore, we could demonstrate that the removal of the mutations had no effect on binding specificity.
这是第一项描述从免疫B细胞来源获得的针对自身抗原蛋白酶3（PR3）的重组人抗体片段的研究。这些自身抗体的检测已被证明对韦格纳肉芽肿的诊断和监测有效。所述抗体片段（单链抗体片段）是从一名系统性自身免疫患者的IgG阳性脾淋巴细胞制备的噬菌体展示文库中分离出来的。克隆策略的设计旨在保持抗体可变基因库的多样性，对几个可变基因的测序表明，所有主要的重链和轻链家族均有代表。我们发现脾淋巴细胞中特定重链可变区的比例过高，这与外周血淋巴细胞中常见的情况不同。经过一轮筛选后，有可能获得针对PR3的特异性单链抗体片段，并且其结合可被患者血清抑制。尽管发现脾库中的抗体片段发生了高度突变，但有趣的是，所选的单链抗体片段仅显示出有限的体细胞突变。此外，我们可以证明去除这些突变对结合特异性没有影响。
