Coupling evaporation-based, microfluidic concentration and confocal fluorescence spectroscopy

Detection limits in confocal fluorescence spectroscopy (CFS) have traditionally been restrained by the low molecular detection efficiencies associated with femtoliter probe volumes. In this report, we address this issue by designing a microfluidic evaporator capable of accepting large sample volumes and concentrating biomolecules to a nanoliter-sized, interrogation chamber. Single molecule fluorescence detection within this chamber is enhanced through microfluidic recirculation, enabling single molecule analysis comparable to traditional capillary-based platforms. Proof of concept is demonstrated using a I OX sample concentrator upstream to a 5 nanoliter CFS detection chamber and recording the subsequent increase in single molecule fluorescent bursts. This marriage of active microfluidics and sample processing, and CFS technology offers a novel means of overcoming the limits of single molecule detection in solution.