Mazza M M, Retegui L A
Instituto de Quimica y Fisicoquimica Biológicas, Buenos Aires, Argentina.
Immunology. 1989 Jun;67(2):148-53.
We re-investigated the properties of a monoclonal antibody (mAb), 4D11, to human growth hormone (hGH) that showed a very weak affinity, recognizing hGH only when the hormone was solubilized on a solid surface. MAb4D11 did not significantly bind 125I-hGH. It was found that three mAb directed to different hGH epitopes (mAb 3C11, 10C1 and NA71) were able to induce the binding of the soluble antigen to mAb 4D11. The co-operative effect could be demonstrated by the formation of binary complexes (Ag:Ab, 1:2) detected by high-performance liquid chromatography (HPLC) and by the increase of radioactivity found when the synergistic mAb were added to 125I-hGH incubated with mAb 4D11 immobilized on polyvinyl microplates. Other possible explanations, such as the formation of cyclic complexes or the generation of a new epitope in the Fc fragment of the first antibody (Ab), were dismissed because the Fab fragment of one of the enhancing mAb (3C11) gave the same effect as the intact Ab. The data suggest that the hGH molecule undergoes a localized conformational change after binding to mAb 3C11, NA71 or 10C1 and that mAb 4D11 binds with high affinity to the modified region of the hormone. The formation or not of ternary complexes (Ag:Ab, 1:3) was used to localize the 4D11 epitope on the surface of the Ag. It is suggested that mAb 4D11 recognizes a conformational change produced in the region defined by the AE5/AC8 epitopes, which is close to the hGH antigenic domain only expressed when the protein is immobilized on plastic surfaces.
我们重新研究了一种针对人生长激素（hGH）的单克隆抗体（mAb）4D11的特性，该抗体亲和力非常弱，仅在激素溶解于固体表面时才能识别hGH。单克隆抗体4D11与¹²⁵I-hGH的结合不明显。研究发现，三种针对不同hGH表位的单克隆抗体（单克隆抗体3C11、10C1和NA71）能够诱导可溶性抗原与单克隆抗体4D11结合。这种协同效应可通过高效液相色谱（HPLC）检测到的二元复合物（抗原：抗体，1:2）的形成以及当将协同单克隆抗体添加到与固定在聚乙烯微孔板上的单克隆抗体4D11孵育的¹²⁵I-hGH中时发现的放射性增加来证明。其他可能的解释，如环状复合物的形成或第一抗体（Ab）的Fc片段中产生新表位，均被排除，因为其中一种增强单克隆抗体（3C11）的Fab片段产生的效果与完整抗体相同。数据表明，hGH分子在与单克隆抗体3C11、NA71或10C1结合后会发生局部构象变化，并且单克隆抗体4D11与激素的修饰区域具有高亲和力结合。三元复合物（抗原：抗体，1:3）的形成与否被用于在抗原表面定位4D11表位。有人提出，单克隆抗体4D11识别由AE5/AC8表位定义的区域中产生的构象变化，该区域靠近仅在蛋白质固定在塑料表面时才表达的hGH抗原结构域。
