Engineering a Proximity Biosensor via Constitutional Dynamic Chemistry

Affinity binding‐induced DNA assembly is a fundamental principle for designing proximity biosensors for sensitive and wash‐free protein detection and imaging. However, current design strategies for these biosensors face an intrinsic trade‐off between binding affinity and background signal. Here, we demonstrate that this intrinsic issue can be addressed by using constitutional dynamic chemistry as a guiding principle in the rational design of proximity biosensors. As exists in a dynamic equilibrium, the constitutional dynamic network (CDN)‐based proximity biosensors can be adjusted to maximize the affinity to the target protein while minimize non‐specific interactions that contribute to background signals. By further detecting the ratio of agonist to antagonist within the CDN, we also significantly improved assay robustness, enabling the sensitive detection of antibodies in complex matrices such as human serum. With the high affinity, low background, and high robustness, we anticipate that our CDN‐based design strategy will find wide applications in biosensor development. Our study also opens the possibility to engineer protein‐responsive synthetic systems with complex dynamic behaviors and function.