We have developed an assay system that can quantitatively determine protein adsorption of different materials down to submonolayer surface coverage at low solution concentrations under flow conditions. Understanding and controlling protein adsorption under these conditions will be a key element in the performance of microdevices that interact with proteins and other biomolecules because the surface area to volume ratio of these devices is substantially larger than those of current systems. Alkaline phosphatase and glucose oxidase were evaluated as probes for quantitative protein adsorption, and loss of protein due to adsorption onto capillary surfaces under flow conditions has been determined at fractional surface coverages on a variety of microfluidic materials. It was found that polyether ether ketone (PEEK) and Teflon had high affinity for both enzymes, while unmodified fused silica, fused silica treated with poly(ethylene glycol), and polyacrylamide had much lower affinity for the two enzymes under the conditions tested. Experimental confirmation of laminar diffusion was also demonstrated with this assay system.