Dueñas M, Malmborg A C, Casalvilla R, Ohlin M, Borrebaeck C A
Department of Immunotechnology, Lund University, Sweden.
Mol Immunol. 1996 Feb;33(3):279-85. doi: 10.1016/0161-5890(95)00137-9.
The display of antibody fragments on the surface of filamentous bacteriophages and the selection of binders from antibody libraries have provided powerful tools to generate human antibodies. We reported recently a new concept (SAP system) for the selection of specific phages by linking antigenic recognition and phage replication, using a soluble fusion protein containing the antigen and a fragment of the M13 coat protein 3. In this investigation, a model library has been composed using six different antibody fragments which were characterized individually regarding their kass, kdiss and Ka. All Fab fragments were specific for a 15 amino acid region of the V3 loop of gp120 (HIV-1). We demonstrated that the SAP system could discriminate between the kinetic parameters of each clone, using different selection strategies. Phages expressing high affinity clones were selected preferentially using low doses of antigen but clones of lower affinity also could be selected by increasing the antigen concentration or using a preselection procedure. Phages expressing antibody fragment with high association or low dissociation rate constants were retrieved by utilizing short contact times between antigen and antibody or antigen-chase conditions.
丝状噬菌体表面抗体片段的展示以及从抗体文库中筛选结合物，为产生人源抗体提供了强大工具。我们最近报道了一种新的概念（SAP系统），用于通过连接抗原识别和噬菌体复制来筛选特异性噬菌体，该系统使用了一种包含抗原和M13外壳蛋白3片段的可溶性融合蛋白。在本研究中，构建了一个模型文库，使用了六个不同的抗体片段，分别对它们的结合速率常数（kass）、解离速率常数（kdiss）和解离常数（Ka）进行了表征。所有Fab片段都对gp120（HIV-1）V3环的一个15个氨基酸区域具有特异性。我们证明，SAP系统可以使用不同的筛选策略区分每个克隆的动力学参数。使用低剂量抗原优先筛选表达高亲和力克隆的噬菌体，但通过增加抗原浓度或使用预选程序也可以筛选出亲和力较低的克隆。通过利用抗原与抗体之间的短接触时间或抗原追赶条件，检索到表达具有高缔合速率常数或低解离速率常数抗体片段的噬菌体。
