Micromachining technology was used to prepare chemical analysis systems on glass chips (1 centimeter by 2 centimeters or larger) that utilize electroosmotic pumping to drive fluid flow and electrophoretic separation to distinguish sample components. Capillaries 1 to 10 centimeters long etched in the glass (cross section, 10 micrometers by 30 micrometers) allow for capillary electrophoresis-based separations of amino acids with up to 75,000 theoretical plates in about 15 seconds, and separations of about 600 plates can be effected within 4 seconds. Sample treatment steps within a manifold of intersecting capillaries were demonstrated for a simple sample dilution process. Manipulation of the applied voltages controlled the directions of fluid flow within the manifold. The principles demonstrated in this study can be used to develop a miniaturized system for sample handling and separation with no moving parts.
Microchips for integrated capillary electrophoresis systems were produced by molding a poly(dimethylsiloxane) (PDMS) silicone elastomer against a microfabricated master. The good adhesion of the PDMS devices on clean planar surfaces allows for a simple and inexpensive generation of networks of sealed microchannels, thus removing the constraints of elaborate bonding procedures. The performance of the devices is demonstrated with both fast separations of φX-174/HaeIII DNA restriction fragments labeled with the intercalating dye YOYO-1 and fluorescently labeled peptides. Detection limits in the order of a few zeptomoles (10(-)(21) mol) have been achieved for each injected DNA fragment, corresponding to a mass detection limit of ∼2 fg for the 603 base pair fragment. Single λ-DNA molecules intercalated with YOYO-1 at a base pair-to-dye ratio of 10:1 could be detected with an uncomplicated laser-induced fluorescence detection setup. High single-molecule detection efficiency (>50%) was achieved under electrophoretically controlled mass transport conditions in PDMS microchannels.
2097Micromachined capillary electrophoresis systems with integrated sample injection have been fabricated on glass chips using standard photolithographic and etching techniques. The injector permits volume-defined electrokinetic sample injection without sample biasing. Utilization of short separation capillaries and high field strengths in combination with a small sample plug length results in both fast and efficient separations of fluorescein isothiocyanate-(FITC-) labeled amino acids. Analysis times range from a few seconds to a few tens of seconds with corresponding plate numbers of 5800-160 000, respectively. Plate heights down to 0.3 pm have been obtained using a separation length of 24 mm and an electric field of 1 kV/cm. As it turns out, a maximum separation efficiency has been reached, limited only by diffusion and the effects of both injection and detection. Automated repetitive sample injection and separation on a time scale of seconds is demonstrated and provides a route to quasi-continuous on-line monitoring of chemical species in a sensorlike fashion.
A micromachined chemical analysis device based on capillary electrophoresis has been successfully used for very fast size separation of a synthetic mixture of fluorescent phosphorothioate oligonucleotides ranging from 10 to 25 bases in length.The device consists of a channel system which has been formed in the surface of a glass plate by a standard photolithographicalprocess. An integrated volume-defined sample injector allowed for unbiased electrokinetic introduction of sample plugs of 150jum length (corresponding sample volume, 90 pL) into the separation channel. This well-defined injection procedure, in combination with the application of high electric fields of up to 2300 V/cm, resulted in size separation of single-stranded oligonucleotides in less than 45 s when a separation distance of 3.8 cm was used. Column efficiencies of up to 200 000 theoretical plates with an associated plate height of 0.2 ^m were obtained. Fast repetitive sample injection and analysis could be demonstrated with excellent reproducibilities for both migration time (<0.06%) and peak height (< 1.7%). The results might also be of relevance for DNA sequencing, where fast oligonucleotide analysis is of key importance. Moreover, they provide a route to "on-line" analysis of synthetic and natural oligonucleotides and possibly other classes of biopolymers.The feasibility of the integration of miniaturized separation techniques into compact devices by using standard photolithographic fabrication procedures has recently been demonstrated in a number of exemplary studies.1-3 In the case of electrical field driven separation techniques, functional models for high-speed capillary electrophoresis (CE),4-7 synchronized cyclic capillary electrophoresis (SCCE),1 2345678 and free flow electrophoresis (FFE)9 have been developed and their separation performance has been evaluated.
Electroosmotic pumping is highly efficient in capillaries of less than 100 mu m inner diameter bearing an immobilized surface charge. Electric fields in the kV cm-1 range allow for liquid motion of several mm s-1 in the case of an aqueous electrolyte. This pumping mechanism is used for miniaturized chemical analysis systems. Flow and mixing behaviour in branched channels are characterized. A capillary electrophoresis device allows for repetitive, electroosmotic injections of 100 pL samples, for efficiencies of up to 200000 theoretical plates in less than a minute, and for external laser induced fluorescence detection at any capillary length of choice between 5 and 50 mm.
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