Cao Peng, Xia Zhinan, Song Wei, Zhang Shuangquan
Jiang su Province Key Laboratory for Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing, 210097 Jiangsu, PR China.
Immunol Lett. 2005 Oct 15;101(1):87-94. doi: 10.1016/j.imlet.2005.05.001.
Elevated levels of B-cell-activating factor of the TNF family (BAFF) have been implicated in the pathogenesis of autoimmune diseases in human. We now report the isolation by phage display of human single-chain antibody fragment (scFv) anti-BAFF. After four rounds of panning against BAFF, thirty-two out of 92 phage clones displayed BAFF binding activity. One of the positive clones, designated F8, bound to BAFF with relatively high affinity and neutralized BAFF bioactivity in vitro. F8 clone was expressed in soluble form in Escherichia coli HB2151 and purified by immobilized metal affinity chromatography (IMAC). The purified scFv recognized BAFF with the affinity constant (K(aff)) of 2.5 x 10(7)M(-1) without cross-reaction to APRIL. In addition to binding, the purified scFv could does-dependently inhibit BAFF-induced mouse spleen B lymphocyte proliferation. Together with its fully human mature, F8 scFv may have therapeutic implications in therapy of autoimmune disorders mediated by BAFF.
肿瘤坏死因子家族的B细胞激活因子（BAFF）水平升高与人类自身免疫性疾病的发病机制有关。我们现在报告通过噬菌体展示技术分离出抗BAFF的人单链抗体片段（scFv）。经过四轮针对BAFF的淘选，92个噬菌体克隆中有32个表现出BAFF结合活性。其中一个阳性克隆，命名为F8，以相对较高的亲和力与BAFF结合，并在体外中和了BAFF的生物活性。F8克隆在大肠杆菌HB2151中以可溶性形式表达，并通过固定化金属亲和色谱（IMAC）进行纯化。纯化后的scFv以2.5×10⁷M⁻¹的亲和常数（K(aff)）识别BAFF，且不与增殖诱导配体（APRIL）发生交叉反应。除了结合作用外，纯化后的scFv还能剂量依赖性地抑制BAFF诱导的小鼠脾脏B淋巴细胞增殖。鉴于其完全人源化的特性，F8 scFv可能在治疗由BAFF介导的自身免疫性疾病方面具有治疗意义。
