We report an insulator-based (or, electrodeless) dielectrophoresis utilizing microfabricated plastic membranes. The membranes with honeycomb-type pores have been fabricated by patterning the SU-8 layer on a substrate which was pretreated with self-assembled monolayer of octadecyltrichlorosilane for the easy release. The fabricated membrane was positioned between two electrodes and alternating current field was applied for the particle trap experiments. The particle could be trapped due to the dielectrophoresis force generated by the non-uniformities of the electric fields applied through the membranes with pores. Simulations using CFD-ACE+(CFD Research, Huntsville, Alabama) suggested that the dielectrophoresis force is stronger in the edge of the pores where the field gradient is highest. The bacteria could be captured on the near edge of the pores when the electric field was turned on and the trapped bacteria could be released when the field was turned off with the release efficiency of more than 93+/-7%. The maximal trapping efficiency of 66+/-7% was obtained under the electric fields (E=128 V/mm and f=300 kHz) when the dilute bacteria solution (Escherichia coli: 9.3 x 10(3) cell/mL, 0.5 mS/m) flowed with a flow rate of 100 microL/min.
We report a practical world-to-chip microfluidic interfacing method with built-in valves suitable for microscale multichamber chip-based assays. One of the primary challenges associated with the successful commercialization of fully integrated microfluidic systems has been the lack of reliable world-to-chip microfluidic interconnections. After sample loading and sealing, leakage tests were conducted at 100 degrees C for 30 min and no detectable leakage flows were found during the test for 100 microchambers. To demonstrate the utility of our world-to-chip microfluidic interface, we designed a microscale PCR chip with four chambers and performed PCR assays. The PCR results yielded a 100% success rate with no contamination or leakage failures. In conclusion, we have introduced a simple and inexpensive microfluidic interfacing system for both sample loading and sealing with no dead volume, no leakage flow and biochemical compatibility.
We have developed a miniaturized bead-beating device to automate nucleic acids extraction from Gram-positive bacteria for molecular diagnostics. The microfluidic device was fabricated by sandwiching a monolithic flexible polydimethylsiloxane (PDMS) membrane between two glass wafers (i.e., glass-PDMS-glass), which acted as an actuator for bead collision via its pneumatic vibration without additional lysis equipment. The Gram-positive bacteria, S. aureus and methicillin-resistant S. aureus, were captured on surface-modified glass beads from 1 mL of initial sample solution and in situ lyzed by bead-beating operation. Then, 10 μL or 20 μL of bacterial DNA solution was eluted and amplified successfully by real-time PCR. It was found that liquid volume fraction played a crucial role in determining the cell lysis efficiency in a confined chamber by facilitating membrane deflection and bead motion. The miniaturized bead-beating operation disrupted most of S. aureus within 3 min, which turned out to be as efficient as the conventional benchtop vortexing machine or the enzyme-based lysis technique. The effective cell concentration was significantly enhanced with the reduction of initial sample volume by 50 or 100 times. Combination of such analyte enrichment and in situ bead-beating lysis provided an excellent PCR detection sensitivity amounting to ca. 46 CFU even for the Gram-positive bacteria. The proposed bead-beating microdevice is potentially useful as a nucleic acid extraction method toward a PCR-based sample-to-answer system.
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