Analytical technologies based on binding assays have evolved substantially since their inception nearly 60 years ago, but our conceptual understanding of molecular recognition has not kept pace. Contemporary technologies, such as single-molecule and digital measurements, have challenged, or even rendered obsolete, core concepts behind conventional binding assay design. Here, we explore the fundamental principles underlying molecular recognition systems, which we consider in terms of signals generated through concentration-dependent shifts in equilibrium. We challenge certain orthodoxies related to binding-based detection assays, including the primary importance of a low dissociation constant (K_D) and the extent to which this parameter constrains dynamic range and limit of detection. Lastly, we identify key principles for designing binding assays that are optimally suited for a given detection application.

基于结合分析的分析技术自近 60 年前问世以来已经有了很大的发展，但我们对分子识别的概念理解并没有跟上。现代技术，如单分子和数字测量，已经挑战了传统结合测定设计背后的核心概念，甚至使其过时。在这里，我们探索了分子识别系统的基本原理，我们根据通过平衡中的浓度依赖性变化产生的信号来考虑这些原理。我们挑战了与基于结合的检测分析相关的某些正统观念，包括低解离常数 ( K _D) 以及该参数限制动态范围和检测限的程度。最后，我们确定了设计最适合给定检测应用的结合测定的关键原则。