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To: tuck who wrote (473)	3/5/2001 1:43:15 AM
From: tuck	Respond to of 1784

Automating Sample Prep for Proteomics . . . Question (circa 1999):>>Microfabricating the World/Chip Fluid Interface for Microfluidic Analytical Systems Prof. D. Jed Harrison, Jed.Harrison@Ualberta.ca Abstract Microfluidic devices offer the power to integrate conventional laboratory methods and entire analysis procedures within a planar, monolithic structure the size of a silicon wafer. Despite the many device examples now in existence illustrating the power of microfluidic technologies, the key issue of the “world to chip” interface must be resolved in order to realistically take advantage of the promise of these systems. Solutions to the problem of the sample-to-chip interface can be broken into two categories. In one mode of operation samples could be introduced to a multi-channel analysis chip using a microtitre-plate-like model. The other mode of operation would involve tubing or capillary connections to fluid solutions in the external environment. Such a design would allow continuous operation for rapid serial analysis of samples, as might be required in a process analytical chemistry environment. At present it is not possible to say which of the above two design concepts will be more feasible for microfluidic devices. The goal of this proposal is to design and fabricate models of both types of interfaces, in order to begin to establish the design constraints microfluidic system designers will operate under when selecting between these two concepts. Potentially useful preliminary designs for both types of multiple-sample inputting interfaces should be produced and evaluated as a result of this effort.<< Answer:>>Integration of immobilized trypsin bead beds for protein digestion within a microfluidic chip incorporating capillary electrophoresis separations and an electrospray mass spectrometry interface. Rapid Commun Mass Spectrom 2000;14(15):1377-83 (ISSN: 0951-4198) Wang C; Oleschuk R; Ouchen F; Li J; Thibault P; Harrison DJ [Find other articles with these Authors] Dept. of Chemistry, University of Alberta, Edmonton, AB T6G 2G2 Canada. A microfluidic device is described in which an electrospray interface to a mass spectrometer is integrated with a capillary electrophoresis channel, an injector and a protein digestion bed on a monolithic substrate. A large channel, 800 microm wide, 150 microm deep and 15 mm long, was created to act as a reactor bed for trypsin immobilized on 40-60 microm diameter beads. Separation was performed in channels etched 10 microm deep, 30 microm wide and about 45 mm long, feeding into a capillary, attached to the chip with a low dead volume coupling, that was 30 mm in length, with a 50 microm i.d. and 180 microm o.d. Sample was pumped through the reactor bed at flow rates between 0.5 and 60 microL/min. The application of this device for rapid digestion, separation and identification of proteins is demonstrated for melittin, cytochrome c and bovine serum albumin (BSA). The rate and efficiency of digestion was related to the flow rate of the substrate solution through the reactor bed. A flow rate of 1 or 0.5 microL/min was found adequate for complete consumption of cytochrome c or BSA, corresponding to a digestion time of 3-6 min at room temperature. Coverage of the amino acid sequence ranged from 92% for cytochrome c to 71% for BSA, with some missed cleavages observed. Melittin was consumed within 5 s. In contrast, a similar extent of digestion of melittin in a cuvet took 10-15 min. The kinetic limitations associated with the rapid digestion of low picomole levels of substrate were minimized using an integrated digestion bed with hydrodynamic flow to provide an increased ratio of trypsin to sample. This chip design thus provides a convenient platform for automated sample processing in proteomics applications. Copyright 2000 John Wiley & Sons, Ltd.<< Dr. Harrison spoke at the Lab Automation conference. Cheers, Tuck