The native and tunable microscale fluid manipulation accessible within 3D-printed configurations can be a transformative tool in biosensing, promoting mass transport and sample mixing to boost assay performance. In this study, we demonstrate that channel height restrictions can support a 2000% acceleration in target recruitment kinetics, a notable 600% improvement in target response magnitude, and a 300% enhancement in assaying selectivity within an entirely reagentless format that requires neither catalytic amplification nor the employment of specialized nanomaterials. This highly accessible experimental configuration supports robust target detection from serum at simple, untreated, and un-passivated sensor surfaces. The underlying operational principles have been elucidated through a combination of theoretical analysis and COMSOL simulation; the enhanced analyte flux leveraged by channel confinement is directly responsible for these effects, which also scale with both bioreceptor surface density and target binding affinity. The operational simplicity of this assaying format with its resolved channel and flux promoted assaying performance, holds significant value not only for biosensing but also for broader microfluidic-integrated applications, such as biosynthesis and biocatalysis.