Determining Binding Kinetics for Microfluidic Carbonylated Protein Enrichment

Microfluidics provides a promising platform for high throughput screening assays and biomolecule sensing through capture. We have recently developed an assay for capturing carbonylated proteins, a common marker of oxidative stress, on a surface-modified polymeric microfluidic chip by taking advantage of carbonyl affinity for hydrazide. Optimization of the biomicrofluidic design involves modeling characteristics inside the chip, including fluid flow and molecular binding constants. We use experimentally calculated concentration changes in a flowing protein solution to determine binding constants between oxidized cytochrome c and oxalyldihydrazide. We assume a simple, bimolecule model that has a first-order interaction. We determine binding constants for association and dissociation between carbonyls and oxalyldihydrazide. These constants will guide our future optimization work for improving carbonylated protein capture on this biomicrodevice. This study is the first to model the binding kinetics between protein carbonyls and oxalyldihydrazide under a flowing condition. These results establish the framework for optimizing microchip design for high-throughput analysis of this common oxidative stress marker, which has been suggested to play a role in the pathogenesis of numerous diseases, including Alzheimer's disease, cardiovascular disease, and diabetes.