Efficient detection of single molecules eluting off an optically trapped microsphere

We demonstrate efficient detection of single fluorescent molecules eluting off a polystyrene microsphere optically trapped in a flowing sheath stream. A 1μm diameter analyte doped microsphere was positioned ∼20 μm upstream of a 16 μm diameter probe laser without significant degradation of the detection signal-to-noise ratio due to scattered laser light and fluorescence from the microsphere. In comparison to more standard capillary sample introduction, the microsphere causes only small perturbations to the sheath fluid flow. The small diameter of the analyte stream eluting from the microsphere results in a greater than 90% detection efficiency for single rhodamine-6G molecules, limited primarily by the photostability of the dye.