A Quantitative First Passage Time Model for Tubular Microfluidic Immunoassays.

Solid-phase immunosorbent reactions, such as ELISA, are widely used for detecting, identifying, and quantifying protein markers. However, traditional centimeter scale well-based immunoreactors suffer from low surface-to-volume (S/V) ratios, leading to large sample consumption and a long assay time. Microfluidic technologies, particularly tubular microfluidic immunoreactors, have emerged as promising alternatives due to their high S/V ratios. Despite experimental advancements, multifactor theoretical studies on tubular microfluidic systems are limited. In this study, we present a theoretical model based on the first passage time method to analyze diffusion-controlled reaction kinetics in tubular microfluidic immunoreactors. We focus on key parameters including binding kinetics, reactor size, and solution viscosity. To validate the model, controlled laboratory experiments were conducted using our in-house developed tip optofluidic immunoassay (TOI). These experimental results confirmed the reliability of theoretical models in the behavior prediction of tubular microfluidic systems under real-world conditions. Our model revealed that accurate and rapid protein biomarker quantification requires not only the development of microscale bioreactors but also the design of next-generation probes with extraordinary binding affinity and specificity. This work offers insights into optimizing critical design parameters in future microfluidic immunoassay development, paving ways for next generation microliter-sized biomolecular analysis.