A 3D finite element model was developed to optimize the kinetics and mass transfer characteristics of low concentration, 18 bp ssDNA targets in bulk media solution, to 18 bp complimentary oligonucleotide probes immobilized on electrochemical detection electrodes positioned along the length of a microfluidic channel. Conditions considered in the model were fluid flow rate, diffusion time, DNA melting temperature, number of matching base pairs, and temperature of the fluid in the channel. System optimization was based on maximizing the uniformity and surface concentration of the specifically bound hybridized DNA, minimizing waste volume generation and the hybridization time. With the coupled simulation method used, the total experiment time was reduced from 150 to 60 min and the simulated results were consistent with experimental results found in the literature. A stopped flow procedure was investigated as a means to improve hybridization. This procedure can not only improve uniformity and capture efficiency, and reduce waste, but can also decrease overall signal intensity relative to continuous flow operation. Finally, the use of temperature in reducing mismatched hybridization and improving duplex stability was also successfully modeled and simulated.