In-line extraction employing functionalized magnetic particles for capillary and microchip electrophoresis.

An approach to performing in-line extraction employing functionalized magnetic particles for CE and microchip electrophoresis is presented. Silica-coated iron oxide particles were synthesized and used as the solid support. The particles were functionalized with octadecylsilane and used as reverse-phase sorbents for in-line SPE followed by electrophoresis. Magnets were used to locally immobilize these sorbents inside the capillary or microchip. Extraction, elution, and detection of the analytes were performed sequentially without interruption or need for sample handling. Mixtures of hydrophobic analytes were successfully extracted from solution using the synthesized magnetic sorbents. CE was able to extract and separate mixture of parabens within 10 min. In-line extraction was also carried out on a disposable PMMA microfluidic device with LIF detection. Electrophoretic separation of fluorescent dyes, Rhodamine 110 and SulfoRhodamine B, was completed in under a minute. The results demonstrated the feasibility of performing the in-line extraction/separation technique in a microchip platform enabling rapid analysis, low sorbent consumption, and increased analyte recovery (relative to the capillary format).

On-chip aptamer-based sandwich assay for thrombin detection employing magnetic beads and quantum dots.

In this paper, we report the development of an on-chip aptamer-based fluorescence assay for protein detection and quantification based on sandwich ELISA principles. Thrombin was selected as a model analyte to validate the assay design, which involves two DNA thrombin aptamers recognizing two different epitopes of the protein. Aptamer-functionalized magnetic beads were utilized to capture the target analyte, while a second aptamer, functionalized with quantum dots, was employed for on-chip detection. The binding of thrombin to the two aptamers via sandwich assay was monitored by fluorescence microscopy. The sandwich assay was performed on disposable microfluidic devices, fabricated on double-sided tapes and polymeric materials using a laser cutting approach. The approach enabled rapid thrombin detection with high specificity. Experimental conditions, such as reagent consumption and incubation time, were optimized in the microchip platform for the lowest limit of detection, highest specificity, and shortest assay time. The analytical performance of the microchip based assay was compared to that in the well plate format (generally utilized for ELISA-based methodologies). The results show that microfluidic chip proved to be a rapid and efficient system for aptamer-based thrombin assays, requiring only minimal (microliter) reagent use. This work demonstrated the successful application of on-chip aptamer-based sandwich assays for detection of target proteins of biomedical importance.

Fabrication of a microfluidic system for capillary electrophoresis using a two-stage embossing technique and solvent welding on poly(methyl methacrylate) with water as a sacrificial layer.

Methods for fabricating poly(methyl methacrylate) microchips using a novel two-stage embossing technique and solvent welding to form microchannels in microfluidic devices are presented. The hot embossing method involves a two-stage process to create the final microchip design. In its simplest form, a mold made of aluminum is fabricated using CNC machining to create the desired microchannel design. In this work, two polymer substrates with different glass transition temperatures (Tg), polyetherimide (PEI) and poly(methyl methacrylate) (PMMA), were used to make the reusable secondary master and the final chip. First, the aluminum mold was used to emboss the PEI, a polymeric substrate with Tg approximately 216 degrees C. The embossed PEI was then used as a secondary mold for embossing PMMA, a polymeric substrate with a lower Tg ( approximately 105 degrees C). The resulting PMMA substrate possessed the same features as those of the aluminum mold. Successful feature transfer from the aluminum mold to the PMMA substrate was verified by profilometry. Bonding of the embossed layer and a blank PMMA layer to generate the microchip was achieved by solvent welding. The embossed piece was first filled with water that formed a solid sacrificial layer when frozen. The ice layer prevented channel deformation when the welding solvent (dichloroethane) was applied between the two chips during bonding. Electrophoretic separations of fluorescent dyes, rhodamine B (Rh B) and fluorescein (FL), were performed on PMMA microchips to demonstrate the feasibility of the fabrication process for microreplication of useful devices for separations. The PMMA micro-chip was tested under an electric field strength of 705 V cm-1. Separations of the test mixture of Rh B and FL generated 55 500 and 66 300 theoretical plates/meter, respectively.