Hand-held microanalytical instrument for chip-based electrophoretic separations of proteins.

The design, fabrication, and demonstration of a hand-held microchip-based analytical instrument for detection and identification of proteins and other biomolecules are reported. The overall system, referred to as muChemLab, has a modular design that provides for reliability and flexibility and that facilitates rapid assembly, fluid and microchip replacement, troubleshooting, and sample analysis. Components include two independent separation modules that incorporate interchangeable fluid cartridges, a 2-cm-square fused-silica microfluidic chip, and a miniature laser-induced fluorescence detection module. A custom O-ring sealed manifold plate connects chip access ports to a fluids cartridge and a syringe injection port and provides sample introduction and world-to-chip interface. Other novel microfluidic connectors include capillary needle fittings for fluidic connection between septum-sealed fluid reservoirs and the manifold housing the chip, enabling rapid chip priming and fluids replacement. Programmable high-voltage power supplies provide bidirectional currents up to 100 microAlpha at 5000 V, enabling real-time current and voltage monitoring and facilitating troubleshooting and methods development. Laser-induced fluorescence detection allows picomolar (10(-11) M) detection sensitivity of fluorescent dyes and nanomolar sensitivity (10(-9) M) for fluorescamine-labeled proteins. Migration time reproducibility was significantly improved when separations were performed under constant current control (0.5-1%) as compared to constant voltage control (2-8%).

Microchip separations of protein biotoxins using an integrated hand-held device.

We report the development of a hand-held instrument capable of performing two simultaneous microchip separations (gel and zone electrophoresis), and demonstrate this instrument for the detection of protein biotoxins. Two orthogonal analysis methods are chosen over a single method in order to improve the probability of positive identification of the biotoxin in an unknown mixture. Separations are performed on a single fused-silica wafer containing two separation channels. The chip is housed in a microfluidic manifold that utilizes o-ring sealed fittings to enable facile and reproducible fluidic connection to the chip. Sample is introduced by syringe injection into a septum-sealed port on the device exterior that connects to a sample loop etched onto the chip. Detection of low nanomolar concentrations of fluorescamine-labeled proteins is achieved using a miniaturized laser-induced fluorescence detection module employing two diode lasers, one per separation channel. Independently controlled miniature high-voltage power supplies enable fully programmable electrokinetic sample injection and analysis. As a demonstration of the portability of this instrument, we evaluated its performance in a laboratory field test at the Defence Science and Technology Laboratory with a series of biotoxin variants. The two separation methods cleanly distinguish between members of a biotoxin test set. Analysis of naturally occurring variants of ricin and two closely related staphylococcal enterotoxins indicates the two methods can be used to readily identify ricin in its different forms and can discriminate between two enterotoxin isoforms.