Figini M, Obici L, Mezzanzanica D, Griffiths A, Colnaghi M I, Winter G, Canevari S
Istituto Nazionale per lo Studio e la Cura dei Tumori, Divisione di Oncologia Sperimentale E, Milan, Italy.
Cancer Res. 1998 Mar 1;58(5):991-6.
The display of repertoires of human antibody (Ab) fragments on filamentous phages and selection by binding of the phage to antigen (Ag) have provided a ready means of deriving human Ab against purified Ag. However, it has been more difficult to obtain phage Ab against an individual Ag of a complex mixture, such as cell surface Ag. Using the technique of "guided selection," we generated human Ab against the high-affinity folate-binding protein (FBP), a cell surface Ag that is overexpressed in many human ovarian carcinomas. The guiding Ab template was provided by the light chain of mouse monoclonal Ab Mov19 (K[aff], 10[8] M[-1]) directed against FBP; this was paired with repertoires of human heavy chains displayed on phages, and the hybrid Ab fragments were selected by binding to an ovarian carcinoma cell line (OVCAR3). The selected human heavy chains were then paired with repertoires of human light chains. Further panning led to the isolation of a human Fab fragment, C4, with a binding affinity of 0.2 x 10(8) M(-1). This was highly specific for FBP, as demonstrated by ELISA and flow cytometry data and by immunoprecipitation of the relevant molecule from the cell surface of ovarian carcinoma cells. Moreover, C4 targeted the same or a closely related epitope of the Ag, as did the template rodent monoclonal Ab Mov19. These results suggest the usefulness of guided selection as a simple means to deriving human Ab against cell surface Ag for which a rodent Ab is available.
在丝状噬菌体上展示人抗体（Ab）片段，并通过噬菌体与抗原（Ag）的结合进行筛选，为获得针对纯化抗原的人抗体提供了一种便捷方法。然而，要获得针对复杂混合物中单个抗原（如细胞表面抗原）的噬菌体抗体则更加困难。利用“导向筛选”技术，我们制备了针对高亲和力叶酸结合蛋白（FBP）的人抗体，FBP是一种在许多人卵巢癌中过度表达的细胞表面抗原。导向抗体模板由针对FBP的小鼠单克隆抗体Mov19（亲和常数K[aff]为10[8] M[-1]）的轻链提供；将其与噬菌体上展示的人重链文库配对，通过与卵巢癌细胞系（OVCAR3）结合来筛选杂交抗体片段。然后将筛选出的人重链与人类轻链文库配对。进一步的淘选导致分离出一种人Fab片段C4，其结合亲和力为0.2×10(8) M(-1)。ELISA、流式细胞术数据以及从卵巢癌细胞表面免疫沉淀相关分子的结果均表明，C4对FBP具有高度特异性。此外，C4靶向的抗原表位与模板啮齿动物单克隆抗体Mov19相同或密切相关。这些结果表明，导向筛选作为一种简单方法，对于获得针对有啮齿动物抗体可用的细胞表面抗原的人抗体是有用的。
