Microfluidic Sequential Injection Analysis in a Short Capillary

Abstract

An automated microfluidic sequential injection analysis system that efficiently manipulates sample and reagent solutions in the nanoliter range in approximately 10 s per analytical cycle is described. The system consisted of a 6-cm-long, typically 75-microm i.d., fused-silica capillary (which functioned as a sampling probe and reactor as well as a flow-through detection cell), a horizontally oriented waste reservoir that provided liquid level differences for inducing gravity-driven flows, an autosampling device holding samples and reagents with horizontally fixed slotted microvials, and a laser-induced fluorescence detection system. Sample and reagent zones were sequentially introduced via gravity-driven flow by scanning the capillary tip (functioning as the sampling probe) through the vial slots, while vials containing sample, reagent, and carrier were sequentially rotated to the probe by programmed movement of the vial holders. Sequentially injected nanoliter zones were rapidly mixed by convection and diffusion within the carrier flow, demonstrating a behavior that conformed well to the Taylor dispersion model, and zone penetration effects were characterized and optimized under Taylor's dispersion theory guidelines. For the determination of fluorescein, a high throughput of 400 h(-1) was achieved, rapidly producing calibration curves (five points) within 45 s. Owing to its adaptability to the Taylor's dispersion model, the system was used also for measuring diffusion coefficients of fluorescent species. Potentials for using the system in enzyme inhibition assays were demonstrated by a reaction involving the conversion of fluorescein digalactoside to fluorescent hydrolysates via beta-galactosidase and the inhibition of beta-galactosidase by diethylenetriaminepentaacetic acid.