Immunoassays are used for routine potency assessment of several vaccines, in some cases having been specifically developed as alternatives to in vivo potency tests. These methods require at least one well characterised monoclonal antibody (mAb) that is specific for the target antigen. In this paper we report the results of the comprehensive characterisation of a panel of mAbs against diphtheria with a view to select antibodies that can be used for development of an in vitro potency immunoassay for diphtheria vaccines. We have assessed binding of the antibodies to native antigen (toxin), detoxified antigen (toxoid), adsorbed antigen and heat-altered antigen. Antibody function was determined by a cell-based toxin neutralisation test and diphtheria toxin-domain recognition was determined by Western blotting. In addition, antibody affinity was measured, and epitope competition analysis was performed to identify pairs of antibodies that could be deployed in a sandwich immunoassay format. Not all characterisation tests provided evidence of “superiority” of one mAb over another, but together the results from all characterisation studies allowed for selection of an antibody pair to be taken forward to assay development.

免疫测定用于几种疫苗的常规效力评估，在某些情况下，已专门开发作为体内效力测试的替代品。这些方法需要至少一种对靶抗原具有特异性的充分表征的单克隆抗体 (mAb)。在本文中，我们报告了一组针对白喉的 mAb 的综合表征结果，以期选择可用于开发体外白喉疫苗的效力免疫测定。我们已经评估了抗体与天然抗原（毒素）、解毒抗原（类毒素）、吸附抗原和热改变抗原的结合。抗体功能由基于细胞的毒素中和试验确定，白喉毒素域识别由蛋白质印迹确定。此外，还测量了抗体亲和力，并进行了表位竞争分析，以鉴定可以部署在夹心免疫测定形式中的抗体对。并非所有表征测试都提供了一种 mAb 优于另一种 mAb 的证据，但所有表征研究的结果一起允许选择抗体对以进行检测开发。