Exploring multivalency-driven sensitivity modulation for optimization and fine-tuning of avidity-based biosensors

The development of multivalent nanoprobes has garnered considerable interest due to their enhanced sensitivity and precision in diagnosing and monitoring diverse diseases. Despite significant advances in nanobiotechnology, the optimal density of binding motifs to maximize the diagnostic efficacy of biosensors remains incompletely understood. Herein, we investigate the influence of multivalency in the functional performance of avidity-based biosensors. By post-translationally modifying the surface of M13 bacteriophages, we achieved multivalent displays of epidermal growth factor receptor-specific repebodies and enhanced green fluorescent proteins in a controlled configuration. Through an array of phage-based nanoprobes, our findings reveal an inverted U-shaped correlation between the degree of multivalency and biosensor sensitivity. Briefly, excessive multivalency negatively impacts analytical performance, likely due to irreversible target depletion, while fine-tuned multivalency significantly enhances antigen recognition and signal amplification. Additionally, the advantages of avidity-demonstrated as improved targeting specificity and sensing capability-are especially prominent in nanoprobes incorporating low-affinity repebodies. This work underscores the critical role of multivalency optimization in biosensor design, providing valuable insights for the development of highly sensitive and reliable diagnostic tools in clinical settings.