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ab-on-a-chip (LOC) devices have garnered a great deal of attention because they offer the possibility of miniaturizing analytical instruments. However, pumping mechanisms within these devices can be very complex and difficult to implement reproducibly. Plastic disks that resemble a compact disc (CD) may provide a simple alternative for pumping within LOCs. Such systems pump fluid using centrifugal force that is generated when the disc spins, eliminating the need for internal moving parts and voltage potentials like those needed for many other LOCs. “I think its simplicity is beautiful,” says Jed Harrison of the University of Alberta (Canada). Interest in CD fluidics for commercial applications actually resulted from frustration with CD-based fluidics chip-based microfluidics. One company, Gamera, which is now part of Tecan, investigated ways to miniaturize their product around eight years ago, but they were unhappy with the “state of may offer a simple the art” in microfluidics, says Greg Kellogg of Tecan. Amersham Biosciences, which spun off their CD technology as Gyros, wanted to stay away from silicon and glass, according to Rolf pumping alternative Ehrnström of Gyros. Centrifugal flow “The real core of the technology, the real basis for it, is the spinning [of the disc] to pump the fluid,” says Kellogg. This centrifugal pumping, says Harrison, “is really only dependent on viscosity and rotation speed, and these are well
Michael J. Felton
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within our current knowledge and comprehenCD valves sion.” In addition, David O’Bryan of Centrifugal analyzers (or clinical analyzers) Burstein Technologies points out that a use centrifugal force as a pump and were You couldn’t take CD-based device is a “natural centrifuge”, developed in the 1960s and 1970s. “But which can be used to process samples. those fluid operations tend to be sort of an arbitrary sample of One benefit of this pumping method coarse, moving stuff from point A to may be that larger samples are more easily point B,” says Kellogg. Although both river water and drop it handled by CD than by electroosmotic flow centrifugal analyzers and CD-based mi(EOF), according to Marc Madou of the crofluidics use centrifugal force, “surface in an EOF system. University of California–Irvine. “100 microforces combined with the smaller dimenliters is large and is easily doable on the CD,” sions afforded by microfabrication allow the he says, suggesting also that CDs may be apformation of passive valves,” he says. As a replied to clinical diagnostics, where larger samples sult, surface chemistry has become one of the priare used. A second benefit over EOF is “that electrokimary methods of forming valves in CDs—an approach netics suffers tremendously from sample matrix effects,” says that does not work with centrifugal analyzers. Harrison. “You have to worry about ionic strength and other Most plastics are extremely hydrophobic, so capillary action variables. It’s fine when you are testing a known sample, but will not occur, says Ehrnström. “You need to make the chanyou couldn’t take an arbitrary sample of river water and drop it nels hydrophilic so the capillary action can work,” he added. in an EOF system.” Other pumping techniques, such as mi- Hydrophobic surfaces, or small apertures, create so-called burst cropumps, show promise but have not proved themselves com- valves to stop fluid flow. The liquid passes through a channel mercially, Harrison adds. and reaches a hydrophobic area, which it will not flow into. At In addition, centrifugal flow is “true parallel analysis,” accord- a higher spin-rate, centrifugal force will overcome the surface ing to Ehrnström. “There are very few other chips that can re- tension and fluid will flow past the hydrophobic patch, says Ehrnström. “In certain cases, when the amount of organic solvent or detergent concentration is very high and will ‘poison’ or destroy the [burst] valve, we use ‘geometric’ valves, which more or less operate like the water lock of a toilet,” says Ehrnström. However, these valves are not air tight; if liquids are stored in the disc, evaporation may occur and contaminate other areas of the disc, according to Madou. ally give reproducible results for many parallel analyses.” The Tecan and Gyros CDs are composed of numerous analysis channels that radiate from the center of the disc toward the outer edge, allowing the centrifugal force to be the same on all the channels. In addition, because the disc is spinning around, “the conditions are essentially identical everywhere,” says Kellogg. There are drawbacks to using centrifugal motion. “A little more thought has to go into the design,” says Harrison. “The only control you have is to spin faster, so the rest has to be hard-wired, and you have to very careful about the arrangement.” Kellogg adds, “You can tweak it a little through operating parameters, but you can’t go from a three-fluid assay to a five-fluid assay with three times the volume.” Other microfluidic technologies are more flexible. “I would like to see devices where you could have the full freedom of moving liquids around,” says Andreas Manz of the Imperial College (U.K.). “I personally think that pumping is not the key issue.” 304 A
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Manufacturing “The CD manufacturing process is reasonably well known and, as such, is available to be adapted to the needs of microfluidics, and I think that is a very good idea,” says Manz. At Gyros, Ehrnström says, this has meant that work on the CDs and instrumentation can happen concurrently, saving time. Tecan has used existing CD drive technology to develop a dual-use CD instrument and detector, which, according to Kellogg, allows people to try out the technology without having to completely commit to it. On the research side, Madou comments, “We have found that it’s extremely easy and fast to make CDs using the soft lithography approach from George Whitesides” (which can also be used for making chip-based microfluidics). In research applications, the discs can be cleaned and used over and over again. “We sometimes just use sticky tape, which you wouldn’t do in the commercial world,” he says. Some commercial researchers use embossing, which is basi-
cally stamping a plastic disc with a master die, to build prototypes to test. Kellogg points out the benefits of embossing: It uses the same plastics as the manufacturing, it can create relatively large numbers of identical discs, and the expense of the embossing equipment has diminished greatly. Commercial manufacturing focuses on disposable CDs, which are made by relatively inexpensive injection molding, as opposed to the expensive silicon etching needed to make some LOCs. In injection molding, the maker injects plastic into a mold to create the disc. “However, this is the easy step; then there are a number of other steps involved in making the final product,” says Ehrnström. It may be necessary to wash the disc, modify the surface chemistry, add chromatographic beads, and attach a lid before it is ready for use. Recently, manufacturing these CD devices got an unexpected boost from the telecom industry due to its financial troubles. Ehrnström says, “We have been able to get hold of very advanced high-tech machines . . . [and] very, very skilled production personnel [who have] worked with this type of manufacturing.” In addition to focusing on the CD disc structure, Burstein has focused on modifying existing CD drive and optics technology. They use these to create a scanning optical microscope that, according to O’Bryan, can view and capture images of microAbove: Multiple samples being pipetted into sized objects such Tecan’s LabCD for analysis. Previous page: as cells on the Gyrolab MALDI SP1 disc for preparing samsurface of a spinples for MALDI analysis. ning disc. The optics measure laser light that is reflected or absorbed, and the system even uses the addressing system, licensed from Phillips, that CDs and DVDs employ to locate areas on the disc.
Applications Drug screening has become the first major use of CD fluidics. In fact, Kellogg says that the employees at Gamera were surprised that the company was being purchased to develop products for drug discovery rather than diagnostics, but “since then, it has made a lot of sense to us.” There are two commercial CD fluidics systems out now—one for drug discovery and one for MALDI sample preparation— and more applications, ranging from diagnostics to PCR, are being researched in industry and academia. The first CD-based product was Tecan’s LabCD, which conducts assays used in adsorption and metabolism studies for drug discovery. LabCD can currently run two different assays, and more are being developed. Kellogg adds that Tecan plans to improve the LabCD discs to use smaller volumes, add new functions, and make it more user friendly. Gyros’ Gyrolab conducts sample preparation of peptides for analysis by MALDI MS. According to Ehrnström, a reversedphase column composed of beads adsorbs the peptides, the beads are washed to remove salt, and then the peptides are eluted using an organic solvent, often acetonitrile, that contains a MALDI matrix. The peptide and matrix solution is slowly eluted to an exposed area of the disc (the rest of the disc is covered by a lid) to allow evaporation of the solvent to cause crystallization. Burstein Technologies’ BCD system, planned for late 2004, will be a medical diagnostics instrument that detects the binding of biological molecules. O’Bryan says that the drive’s optics “can quantitatively count objects bound to the disc, measure optical density changes, and detect size and shape changes with a resolution of about 2 microns.” The first application will be the determination of blood type (ABO/Ph) in about 10 min by using real-time recognition of cells. A disc that measures several cardiac risk factors in a patient’s blood is planned to follow this. BCD will use CD/DVD data storage techniques to let each disc carry instructions explaining how it is to be used. For fluid handling, “many of the current disc designs have very J U LY 1 , 2 0 0 3 / A N A LY T I C A L C H E M I S T R Y
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creates a depth-of-focus problem because simple microfluidics that don’t require the path length through sample wells features like burst valves,” O’Bryan You could say by on the CD will be very short. In adsays, but additional discs with more dition, O’Bryan says that Burstein’s sophisticated microfluidics are being now that we have in our BCD sensitivity might be limited designed. due to the small sample size conAcademic researchers are also purdesign toolbox, 15, 20, or tained on the disc. suing medical diagnostics. Madou, for example, says he is “mainly maybe more common elements looking at sample preparation, cell The next step lyses, and eventually going all the In 2003, Gyros plans to introduce a that we keep on playing with way from blood to DNA.” To fursecond CD that will use affinity chrother medical diagnostics applications, matography instead of a reversedin different variations. he is working on methods to store phase column. The new column will use chemicals on the disc so that they can be streptavidin-coated beads and will bind used easily in the field. One method of storbiotinylated agents for the user’s choice of age is lyophilizing, or freeze-drying, which can antibodies; the bound molecules will be debe used with reagents that can later be rewetted. tected by fluorescence. However, Madou says, “Not everything can be dried, and Due to the growing number of fluid analytical processes so we are looking for a combination of certain reagents lyophilized that can be included on the discs, many uses for CD-based sysand other ones not.” Due to the types of valves currently used tems are being contemplated. “You could say by now that we on CDs, liquids stored on the CD may evaporate and/or con- have in our design toolbox, 15, 20, or maybe more common taminate other areas of the disc. Madou is working on physical elements that we keep on playing with in different variations,” valves that will block vapor as well as liquid. says Madou. “Interested parties come to us and say, ‘Look I Madou and others are also researching detectors on the disc, have this assay, and it has these steps,’ . . . Do we have these and he says, “We were the first ones to do a two-point calisteps available on the CD?” Gyros has bration of an optode on a CD.” grown cells on the disc and worked He adds that detection with cell-based assays and genomics, on the disc and Tecan has examined handling large sample volumes and isolating small amounts of DNA from them. O’Bryan says that Burstein has “also looked at applications related to food treatment, forensic analysis, veterinary testing, biosafety, pharmaceutical clinical trials, and proteomics.” One area that CD makers are looking at is PCR and applications that are now served by chip-based systems. Gyros and Tecan both use polyolefinbased plastics that can handle being thermocycled for PCR. In the end, CD fluidics may prove to be a simple answer to miniaturization in some situations. “I wouldn’t be surprised if they did as well as Caliper does with voltage drive and Affymetrix does with a very primitive approach to fluidics but a sophisticated glass surface,” says Manz. We will have to wait and see how successful this “beautiful simplicity” will be.
Microchannels, valves, and reaction chambers on Gyros’ Gyrolab microlaboratory disc. 306 A
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Michael J. Felton is an associate editor of Analytical Chemistry.
