The measurement of proteins with a limited number of available non-overlapping epitopes recognizable by antibodies represents a common challenge for the development of drug-tolerant clinical biomarker assays. For target proteins with two dominant epitopes, only one epitope remains when the other is occupied by the therapeutic antibody. Alternative strategies for overcoming this obstacle have been described in the literature; however, these methods have potential limitations. We have developed a novel method for measuring target engagement when only one epitope remains after therapeutic antibodies bind their analytes. The method combines Affinity Capture Elution (ACE) followed by simultaneous capture and detection of the protein of interest. This novel method has been named ACE-Sandwich. The application of this method is not dependent on the immunoglobulin G subclass of the therapeutic antibody, nor does this method require sample pretreatment. Furthermore, the ACE-Sandwich method is highly sensitive, reproducible, and tolerant to high concentrations of therapeutic antibody.

具有有限数量的可被抗体识别的可用非重叠表位的蛋白质的测量代表了耐药性临床生物标志物检测开发的共同挑战。对于具有两个显性表位的靶蛋白，当另一个表位被治疗性抗体占据时，仅保留一个表位。克服这一障碍的替代策略已在文献中有所描述；然而，这些方法有潜在的局限性。当治疗性抗体与其分析物结合后只剩下一个表位时，我们开发了一种测量靶标参与度的新方法。该方法结合了亲和捕获洗脱 (ACE)，然后同时捕获和检测感兴趣的蛋白质。这种新颖的方法被命名为ACE-Sandwich。这种方法的应用不依赖于治疗性抗体的免疫球蛋白 G 亚类，也不需要样品预处理。此外，ACE-Sandwich 方法对高浓度的治疗性抗体具有高度敏感性、可重复性和耐受性。