Immunoassays have become much more sophisticated since the enzyme linked immunoassays became widely used. Microfluidics in particular, coupled with advanced optical and electrochemical readout systems have reduced the limits of detection, decreased assay time, and simplified automation. Yet the sensitivity of the microfluidic immunoassays is still limited by the ability of the detector to discriminate between signal and background. Three main approaches to produce higher signal/background are reviewed and critiqued: target preconcentration, reaction confinement in a small detection volume and signal amplification strategies. Combinations of these strategies can be used to increase sensitivity and may provide clinical diagnostics for biomarkers present in very low concentrations.