Neri D, Petrul H, Winter G, Light Y, Marais R, Britton K E, Creighton A M
Cambridge Centre for Protein Engineering-MRC Centre, UK.
Nat Biotechnol. 1996 Apr;14(4):485-90. doi: 10.1038/nbt0496-485.
A wide range of antibody fragments can be expressed in bacteria and detected immunochemically via peptide tags. Using specially designed tags, we have developed a strategy for radiolabeling antibody fragments secreted from bacteria. Tagged antibody fragments were secreted either into the bacterial periplasm or the culture medium. The tag was not subject to proteolysis either in the broth or in human plasma. After affinity purification the antibody fragments were phosphorylated with [gamma-32P]ATP and casein kinase II. The labeled fragments were used in a gel band-shift assay to measure antigen binding affinities. In contrast to non site-specific methods such as radioiodination, antibodies labeled with casein kinase II retain full immunoreactivity. Radioactively phosphorylated antibody fragments may have many other applications, including radioimmunoassays and radioimmunotherapy.
多种抗体片段可在细菌中表达，并通过肽标签进行免疫化学检测。利用专门设计的标签，我们开发了一种对细菌分泌的抗体片段进行放射性标记的策略。带标签的抗体片段可分泌到细菌周质或培养基中。该标签在肉汤或人血浆中均不会被蛋白水解。亲和纯化后，抗体片段用[γ-32P]ATP和酪蛋白激酶II进行磷酸化。标记的片段用于凝胶迁移试验以测量抗原结合亲和力。与放射性碘化等非位点特异性方法不同，用酪蛋白激酶II标记的抗体保留了完整的免疫反应性。放射性磷酸化的抗体片段可能还有许多其他应用，包括放射免疫分析和放射免疫治疗。
