Flow Cytometry, Beads, and Microchannels

Microfluidic devices generally consume microliter to submicroliter volumes of sample and are thus well suited for use when the required reagents are scarce or expensive. Because microfluidic devices operate in a regime in which small Reynolds numbers govern the delivery of fluid samples, reagent mixing and subsequent reactivity has been a severe limiting factor in their applicability. The inclusion of packed beads in the microfluidic device repertoire has several advantages: molecular assemblies for the assay are created outside the channel on beads and characterized with flow cytometry, uniform populations of beads may be assured through rapid cytometric sorting, beads present a larger surface area for the display of receptors than flat surfaces, rapid mixing in the microcolumn is achieved because the distance that must be covered by diffusion is limited to the (≤1-μm) interstitial space between the closely packed receptorbearing beads, and analytes are captured in a flow-through format and, as such, each bead can act as a local concentrator of analytes. Progress in the combined use of beads and microfluidic devices has been limited by the ability to pack beads at specific sections of microfluidic devices. A subsequent challenge associated with the packed microcolumns of beads is the substantial pressure drop that affects the fluid flow velocity. However, some of these challenges have been overcome in the design of simple model systems that have potential applications in DNA analysis, chromatography, and immunoassays. It is the intent of this chapter to examine the recent emergence of small-volume heterogeneous immunoassays, using beads trapped in microchannels, while excluding other closely related applications such as capillary electrophoresis and flow injection–based approaches. Of necessity, the authors’ interests and availability of information pertinent to the specific discussions presented below impose additional restrictions. To date, there are only a handful of applications that combine packed beads and microfluidic devices, and even fewer that make the overt connection between flow cytometry–based assays and beads. Harrison and coworkers have provided one of the earliest conceptual demonstrations of the capability to incorporate packed beads in microfluidic devices for analytical purposes.