Microfluidic biopanning can reduce a sample size and improve the reproducibility of biopanning. However, there is no practical guideline for optimal experimental conditions to identify high affinity peptides, including incubation and washing times and washing and elution flow rates. Here we investigated the elution dynamics of M13 phage displaying a well-known streptavidin-binding peptide motif, histidine-proline-glutamine (HPQ), to streptavidin immobilized in a polydimethylsiloxane microfluidic channel via a biotin-polyethylene glycol linker. The phages bound to streptavidin were eluted using an acidic buffer, which weakens the molecular interactions of HPQ and streptavidin. Various washing flow rates were examined to determine the effects of drag forces on the elution of phages, and the total force per phage combined with the drag force was calculated. The critical washing drag force and washing time were 5.9 x 10(-8) dyne and 1.1 min, respectively, as obtained by fitting the data using a sigmoidal function. The dissociation rate constant of HPQ-displaying phage (2.0 x 10(-2) min(-1)) was also obtained from the double exponential decay fitting. Our work provides a practically useful approach to the optimization of experimental parameters for the design of a microfluidic biopanning platform.