Compact Disc Microfluidics
CD Microfluidics is a sub-field of microfluidics that deals with the behavior, precise control and manipulated of fluids in the micro-domain. CD Microfluidics takes advantage of centrifugal forces for fluid propulsion. Fluidic channels and reservoirs are embedded in a CD-like plastic substrate and the whole platform is spun on a motor in order to manipulate fluids. A whole range of fluidic functions have been designed and implemented which include valving, decanting, calibration, mixing, metering, sample splitting, and separation. Those fluidic functions have been combined with analytical measurement techniques, such as optical imaging, absorbance, and fluorescence spectroscopy and mass spectrometry, to make the centrifugal platform a powerful solution for medical and clinical diagnostics and high throughput screening (HTS) in drug discovery. Applications of a compact disc (CD) based centrifuge platform include two-point calibration of an optode-based ion sensor, an automated immunoassay platform, multiple parallel screening assays, and cellular-based assays.
Advantages of the microfluidic CD
In order to perform complex fluidic manipulations, required for Lab on CD, our group has developed an optimized various capabilities, including valving (capillary valve, Coriolis valve, syphon valve, serial syphon valve), reciprocal mixing, separation (such as DEP on the CD). These technologies are described below.
Valving Technologies (capillary, hydrophobic, siphon)
Valving is a most important function in any type of fluidic platform. Both hydrophobic and capillary valves have been integrated into the CD platform. Hydrophobic valves feature an abrupt decrease in the hydrophobic channel cross-section, i.e., a hydrophobic surface prevents further fluid flow. In contrast, in capillary valves, liquid flow is stopped by a capillary pressure barrier at junctions where the channel diameter suddenly expands.
In hydrophobic valving, for liquid to move beyond these pressure barriers, the CD must be rotated above a critical speed, at which point the centripetal forces exerted on the liquid column overcome the pressure needed to move past the valve.
Capillary valves have been implemented frequently on CD fluidic platforms. The physical principle involved is based on the surface tension, which develops when the cross section of a hydrophilic capillary expands abruptly.
One of the main challenges to integration for sample-to-answer microfluidic systems is the requirement for serial valving to allow the sequential release of fluids in a temporally and spatially controlled manner. The advantages offered by centrifugal microfluidic platforms make them excellent candidates for integration of biological analysis steps, yet they are limited by the lack of robust serial valving technologies. This is especially true for the majority of centrifugal microfluidic devices that rely on hydrophilic surfaces, where few passive serial valving techniques function reliably. Building on the useful functionality of centrifugal microfluidic siphoning previously shown, a novel serial siphon valve is introduced that relies on multiple, inline siphons to provide for a better controlled, sequential release of fluids.
The serial siphon is shown to be robust and reproducible, with variability caused by the dependence on contact angle, rotation velocity, and fluidic properties (viz., surface tension) significantly reduced compared to current microfluidic, centrifugal serial valving technologies. This design withholds fluids through multiple high and low speed operations and allows the fluids to be subsequently distributed to areas of interest. The serial siphon valve has numerous potential applications, and will be especially useful in the development of IVD systems for NA analysis. Integrated microfluidic NA diagnostic devices can be quite complex, and the avail- ability of a simple and passive serial valve will eliminate the need for many of the more complex valving solutions.
Reciprocating Centrifugal Microfluidics Mixer
Proper mixing of reagents is of paramount importance for an efficient chemical reaction. While on a large scale there are many good solutions for quantitative mixing of reagents, as of today, efficient and inexpensive fluid mixing in the nanoliter and microliter volume range is still a challenge. Complete mixing is of special importance in any small-scale analytical application where the scarcity of analytes and the low volume of the reagents demand that all of the available analytes and reagents are efficiently used. We have demonstrated the design and fabrication of a novel centrifugal force-based unit for fast mixing of fluids in the nanoliter to microliter volume range (Noroozi et.al, Review of Scientific Instruments, vol. 80, no. 7, pp. 075102-8(2009)). The device consists of a number of chambers (including two loading chambers, one pressure chamber and one mixing chamber) that are connected through a network of microchannels, and is made by bonding a slab of polydimethylsiloxane (PDMS) to a glass slide. The PDMS slab was cast using a SU-8 master mold fabricated by a two-level photolithography process. This microfluidic mixer exploits centrifugal force and pneumatic pressure to reciprocate the flow of fluid samples in order to minimize the amount of sample and the time of mixing. Our results suggest that quantitative mixing was achieved in less than three min. This device can be employed as a stand-alone mixing unit or may be integrated into a disk-based microfluidic system where, in addition to mixing, several other sample preparation steps may be included.
[go to] Z. Noroozi, et al., "Reciprocating flow-based centrifugal microfluidics mixer," Review of Scientific Instruments, vol. 80, p. 075102, 2009.
Tissue homogenization and cell lysis are the first steps in virtually all molecular biology and molecular diagnostic techniques. The purpose of cell lysis is to disrupt the cells and release genomic and proteomic material to enable downstream processing. Cell lysis and subsequent nucleic acid or protein purification represent a considerable obstacle commonly encountered in molecular biology sample preparation protocols. Although easily attainable with certain cells and microorganisms, many cells of interest pose significant challenges due to increased cell wall structural integrity. Molecular biology sample preparation kits and equipment are part of a rapidly growing multi-billion dollar market. It has been estimated that up to 15% of researchers’ time is consumed with the simple and redundant task of sample preparation.
The most efficient and elegant method of mechanical cell lysis is bead-beating. Bead-beating is accomplished by combining a liquid or solid sample with milling beads in a closed container and exposing the entire mixture of grinding media, buffer and tissue to intense mixing by rapid and abrupt motion of the container. If a component of angular motion is introduced into a bead-beating system, then additional lysis results from impaction and friction between cells and the lysing matrix as a result of the Coriolis effect. The lysing matrix directly contributes to lysis again during centrifugation, as observed previously in a centrifugal CD-based homogenizer.
No currently commercially available sample lysis and preparation system can handle samples in the microliters volume range. The novel magnetohydrodynamic disk-based homogenization system described herein successfully combines microfluidics, magnetically actuated cell lysis, and lysate clarification by centrifugation in a single device. The device consists of a simple rotary magnetic field superimposed on a rotor-based, disposable CD-like disk containing sample chambers preloaded with lysing matrix and a ferromagnetic blade. The device yields clarified lysates from even the most difficult to lyse microorganisms utilizing up to 70 ’L of substrate. Thus, the magnetohydrodynamic disk-based homogenization system is a feasible solution for efficient lysis on a microscale. Based on results with E. coli and S. cerevisiae we demonstrate that this system can be utilized for semi-automated molecular biology sample preparation on a micro and nano scale sample volume to provide genomic materials. The CD-like magnetically actuated centrifugal lysis system described herein has the additional advantage of facile integration with other CD- based microfluidics analytical systems, and as such promotes the novel concept of an entire molecular diagnostics microfluidics laboratory on a disk.
Separation using Dielectrophoresis on the CD
Dielectrophoresis (DEP) enables the selective manipulation of a targeted particle, or population of particles, using the interaction of a non-uniform electric field with the induced effective dipole moment of the particle(s). DEP is advantageous over other particle separation techniques such as FACS (Fluorescence-activated cell sorting) and MACSÒ (Magnetic-activated cell sorting) because discrimination between different particles is based solely on their intrinsic physical properties. These physical properties determine the particle’s dielectric properties and give it a characteristic dielectric phenotype. The use of DEP eliminates functionalized magnetic beads or fluorophores required by other separation techniques such as magnetophoresis and flow cytometry. The elimination of these often expensive labels reduces assay complexity, time and costs which can expand the availability of diagnostic tests, such as for HIV, and make therapies for cancer or degenerative diseases available to a broader number of patients.
The use of centrifugal forces for fluid pumping and fluid manipulation improves traditional DEP platforms in terms of footprint, cost, robustness and practicality. 3D carbon-electrode DEP, or carbon-DEP, offers key advantages over the use of other DEP techniques including a simple and inexpensive fabrication process. The implemented modularity allows for the use of interchangeable disposable chips depending on the type of assay or study to be conducted. Other electrokinetic-based applications besides filtering are targeted including cell, microorganism and biomolecule sorting and cell electroporation for DNA release and transfection.
[go to] R. Martinez-Duarte, et al., "The integration of 3D carbon-electrode dielectrophoresis on a CD-like centrifugal microfluidic platform," Lab on a Chip, vol. 10, pp. 1030-1043, 2010.