A trap-and-release integrated microfluidic systemfor dynamic microarray applicationsWei-Heong Tan* and Shoji Takeuchi*†‡

*Center for International Research on MicroMechatronics, University of Tokyo, Institute of Industrial Science, 4-6-1 Komaba, Meguro-ku, Tokyo153-8505, Japan; and †Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, 4-1-8 Honcho,Kawaguchi, Saitama 332-0012, Japan

Edited by Richard A. Mathies, University of California, Berkeley, CA, and accepted by the Editorial Board November 30, 2006 (received for reviewAugust 2, 2006).

Dynamic microarrays hold great promise for advancing research inproteomics, diagnostics and drug discovery. However, this poten-tial has yet to be fully realized due to the lack of reliable multi-functional platforms to transport and immobilize particles, infusereagents, observe the reaction, and retrieve selected particles. Weachieved all these functions in a single integrated device throughthe combination of hydrodynamic and optical approaches. Hydro-dynamic forces allow simultaneous transportation and immobili-zation of large number of particles, whereas optical-based micro-bubble technique for bead retrieval gives dexterity in handlingindividual particles without complicated circuitry. Based on thecriterion derived in this paper, the device was designed, andfabricated using standard photolithography and soft lithographymethods. We examined the dynamics of bubble formation anddissipation in the device, and parametric studies revealed thathigher power settings at short intervals were more efficient thanlow power settings at longer intervals for bead retrieval. We alsodemonstrated the capabilities of our device and its potential as atool for screening methods such as the ‘‘one-bead-one-compound’’(OBOC) combinatorial library method. Although both approaches,hydrodynamic confinement and optical-based microbubbles, arepresented in one device, they can also be separately used for otherapplications in microchip devices.

high throughput screening � lab-on-a-chip � MEMS � bead-based assay �microbubble

M icroarray applications are extensive, and have been suc-cessfully used in basic scientific studies (1–4), drug-

discovery (5, 6), and diagnostic purposes (7). Microarrays can bebroadly classified into two categories: static and dynamic. Instatic microarrays, biomolecules and chemicals are immobilizedas microspots on a static solid support, and can be fabricatedusing a variety of technologies, including printing with high-speed arrayer onto glass slides (7), photolithography usingpremade masks (8), or photolithography using micromirrordevices (9). In the case of dynamic microarrays, instead ofstationary solid supports, bio-molecules and chemicals are im-mobilized onto mobile substrates, usually microbeads (10, 11).Besides bead-based microarrays, dynamic microarrays also in-clude cell-based arrays (12, 13). Compared with their staticcounterparts, dynamic microarrays have several advantages: (i)ability to mix-and-match the beads (or cells) to cater for the typeof screening to be performed, and introduce them into themicroarray on demand offer great versatility; (ii) beads (or cells)can be replaced, resulting in a reusable format that greatlyreduces the cost of operation; (iii) reaction on beads tends to befaster compared with conventional planar surfaces, as mi-crobeads have increased surface area, thus higher binding ca-pacity. To fully realize the potential of dynamic microarrays, itwill be necessary to build a platform that allows us to transportparticles, to immobilize them for convenient signal detection, todeliver reagents to them while under continual observation, andto retrieve selected particles. However, despite the progress in

particle handling techniques, there is still no reliable, inexpen-sive and robust device that can perform these functions.

Here, we propose a microchannel design that accomplishes allthese functions (transportation, immobilization, infusion of re-agents, observation, and retrieval) in a single integrated device (Fig.1), and apply it to dynamic microarray applications. The principlebehind this design is simple: Fluidic resistance along the straightchannel is smaller, and beads in the flow will be carried into thetrap; but once the traps are filled, the flow will be redirected to theloop channels. A few groups (14–16) have also reported usinghydrodynamic forces to immobilize beads/cells, but none managedto handle and release selected particles from their devices. More-over, our design criterion allows one to design the device withoutany trial and error. We can achieve one-bead-to-one-trap in thearray and the characteristic of the flow allows us to retrieve atrapped particle from the array by displacing it back into the mainflow. This retrieval of selected beads from the array is achieved bymicrobubbles. The microbubble technique is ideal for our applica-tion compared with electrokinetic (17–19), optical (20), and othertechniques (21) because the force exerted by an expanding bubbleis enough to counter the flow. However, conventional microbubble-based devices (22–24) suffer the drawback of complicated circuitsand connections. In this device, the microbubbles are formed byusing a new optical technique that does away with the need ofcomplicated on-chip integrated circuits and connection to externalcontrol electronics, greatly simplifying fabrication and operation.Consequently, our trap-and-release device can be easily imple-mented with conventional and soft lithography methods. Thecombination of hydrodynamic and optical approaches allows us toeliminate the shortcomings of each method; hydrodynamic ap-proach allows simultaneous manipulation of large number ofparticles, whereas optical-based technique for retrieval gives dex-terity in accessing individual particles.

As a demonstration, we illustrated how this device might beused as a tool for screening methods such as the ‘‘one-bead-one-compound’’ (OBOC) combinatorial library method (2, 4, 10):beads were introduced and arrayed in the device, analyte wasperfused over the beads, and selected positive beads wereretrieved from the device. The device and methodology pre-sented here lay the groundwork for highly parallel economicalbead-based chemical microarrays, and with modifications, itmight even be applicable for cells. We also believe that thetechniques, hydrodynamic confinement and optical-based mi-

Author contributions: W.-H.T. and S.T. designed research; W.-H.T. performed research;W.-H.T. analyzed data; and W.-H.T. and S.T. wrote the paper.

The authors declare no conflict of interest.

This article is a PNAS direct submission. R.A.M. is a guest editor invited by the EditorialBoard.

Abbreviation: OBOC, one-bead-one-compound.

‡To whom correspondence should be addressed. E-mail: takeuchi@iis.u-tokyo.ac.jp.

This article contains supporting information online at www.pnas.org/cgi/content/full/0606625104/DC1.

© 2007 by The National Academy of Sciences of the USA

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crobubbles, can be separately used for other applications inmicrochip devices.

Concept and Design CriterionA Simple Hydrodynamic Trap (�-Fluidic Trap). Fig. 1 A shows theschematic of the �-Fluidic trap. It composes of square-waveshaped loop channels superimposed onto a straight channel, withnarrowed regions along the straight channel functioning as traps.The channels are designed such that when a trap is empty, thestraight channel has a lower flow resistance than that of the loopchannel. As a result, we have bulk of the fluid flowing along thestraight channel. A particle in the flow will be carried by thismain stream into the trap (trapping mode). This particle acts asa plug, increasing the flow resistance drastically along thestraight channel, and redirecting the main flow to the loopchannel. Subsequent particles will then be carried along the loopchannel, bypassing the filled trap (bypassing mode). Based on asimple model, the design criterion for this trap will be derived.

Pressure drop in a microchannel. Using the Darcy–Weisbach equa-tion to determine the pressure drop or pressure difference in amicrochannel and solving the continuity and momentum equa-tions for the Hagen–Poiseuille flow problem, we obtain thepressure difference �p � ƒL�V2/2D, where ƒ is the Darcy frictionfactor, L is the length of the channel, � is the fluid density, V isthe average velocity of the fluid, and D is the hydraulic diameter,respectively. D can be further expressed as 4A/P for a rectangularchannel, and V as Q/A, where A and P are the cross-sectional areaand perimeter of the channel, and Q is the volumetric f low rate.The Darcy friction factor, ƒ, is related to aspect ratio, �, andReynolds number, Re � �VD/�, where � is the fluid viscosity.The aspect ratio is defined as either height/width or width/heightsuch that 0� � �1. The product of the Darcy friction factor andReynolds number is a constant that depends on the aspect ratio,

i.e., ƒ � Re � C(�), where C(�) denotes a constant that is afunction of � [refer to supporting information (SI) Table 2 forsolutions of C(�) (25)]. After simplifications, we obtain theexpression

�p �C(�)

32��LQP2

A3 . [1]

Design criterion for �-Fluidic trap. In Fig. 1 A, we have the simplifiedcircuit diagram of the trap. Fluid can flow from junction A to Bvia path 1 or 2. Ignoring minor losses due to bends, widening/narrowing, etc., Eq. 1 is applied separately for paths 1 and 2, andbecause the pressure drop is the same for both paths, we equateboth expressions to yield

Q1

Q2� �C2(�2)

C1(�1)���L2

L1���P2

P1�2

��A1

A2�3

, [2]

where subscripts 1 and 2 denote paths 1 and 2, respectively. Forpath 1, the length, L1, is assumed to be that of the narrow channelto simplify analysis. This is valid because most of the pressuredrop occurs along the narrow channel. For the trap to work, thevolumetric f low rate along path 1 must be greater than that ofpath 2, i.e., Q1 � Q2. Using the relationships A � W � H and P �2(W � H), where H is the height of the channels, we arrive at

�C2��2�

C1��1����L2

L1� � �W2 � H

W1 � H�2

� �W1

W2�3

� 1. [3]

Note that this final expression does not contain any fluid velocityterm, implying that a properly designed trap will work for allvelocities in the laminar flow regime.

Optical-Based Microbubble Retrieval System. In our �-Fluidic trapdevice, once all of the traps are occupied, the main flow will beredirected to the loop channels. Subsequent particles, not beingable to enter occupied traps, will follow the main flow out of thedevice. Taking advantage of this characteristic, we can retrievea trapped particle from the array by displacing it back into themain flow using microbubbles. Here, we propose a simpleoptical-based method to create microbubbles without any needfor circuits and connections. Fig. 1B shows the schematic of ourmethod. Aluminum patterns, functioning as heaters, are locatednear the narrowed region of the �-Fluidic traps. When we focusan IR laser onto the aluminum pattern, localized heating resultsin bubble formation and the expanding bubble displaces theimmobilized particle from the �-Fluidic trap into the main flow.The displaced particle is then carried by the flow out of thedevice where it can be collected. Size of the bubble can becontrolled by varying the laser power and duration of the appliedlaser.

The schematic of the whole system is depicted in Fig. 1C. Thedevice is mounted onto an inverted microscope with an auto-matic XY stage, which is controlled with a manipulator joystick.Other controllers regulate the intensity and duration of the laser,which is focused through the objective lens. The infusion system(pumps), laser system, and manipulation system can all becontrolled by computer, allowing total automation of the systemin the future.

Results and DiscussionWe connected the �-Fluidic traps in series to create an array forhigh density immobilization of beads. Two devices, one that metand one that did not meet the design criterion (Eq. 3), werefabricated and tested. Table 1 lists the dimensions of bothdesigns. By superimposing time-lapsed images taken with ahigh-speed camera, we retraced the paths taken by beads in theflow for both devices. In device A, Q1/Q2 had a value smaller

Fig. 1. Trap-and-release mechanism and experimental setup. (A) Schematicdiagram of the �-Fluidic trap. When the trap is empty, flow resistance alongthe straight channel is lower than that of the loop channel, and the mainstream flows along the straight channel. A bead in the flow is carried by themain stream into the trap if it is empty (trapping mode). Beads will be carriedalong the loop channel if the trap is filled, bypassing the occupied trap(bypassing mode). This design allows for one-bead-to-one-trap. (B) Releasemechanism using microbubble. IR laser is focused onto the aluminum pattern,causing localized heating and bubble formation. The formed bubble displacesthe trapped particle from the trap into the main flow. The particle is thencarried by the main flow out of the device. (C) Experimental setup.

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than 1, indicating that the bulk of the flow was along the loopchannel. As expected, none of the beads in Device A wasimmobilized in the �-Fluidic traps; all of the beads followed themain flow, zigzagging through the device without entering thetraps (Fig. 2A, see SI Movie 1). On the other hand, Q1/Q2equaled 3.95 for device B. With �80% of the flow along thestraight channel, all of the beads were sequentially carried intothe traps and immobilized (Fig. 2B, see SI Movie 2). Arrows inFig. 2B show the bypassing and trapping mode, respectively. Onehundred microbeads can be arrayed in 20 s.

Our proposed design for hydrodynamic confinement is basedon the principle of fluidic resistance. This passive trappingmechanism is robust and insensitive to fluctuations caused byoccasional gas bubbles introduced during the switching of solu-tions. It is also extremely efficient, thus highly suitable forhandling small samples. Compared with previously reportedhydrodynamic traps (14–16), fabrication is extremely simple, andthe design criterion allows one to design the device without anytrial and error. When designing the trap, height of the micro-channel, H, is an important parameter to consider. It should beset to a value larger than that of the target particle’s diameter(�p). However, if H is too large, the excess leakage after aparticle occupies a trap might result in multiple particles in asingle trap. Based on our experience, H should be set to �p H 1.4 �p for the �-Fluidic trap to achieve one-bead-to-one-trap. Our device is designed to trap particles of a specific size,and we have worked successfully with beads having a coefficientof variation of �4%. Using a mixture of beads that exhibits highpolydispersity might result in suboptimal performance due to thetrapping of multiple beads per site. With the same designcriterion, we have also fabricated a high-density (1 � 104) devicefor immobilization of beads (see SI Text and SI Fig. 5).

The actual trap-and-release device used in our experiments isshown in Fig. 3A. This device is designed to immobilize 100beads, and has traps that are numbered for individual address-

ability. To demonstrate the individual addressability of the beadmicroarray, and the ease of operation of our trap-and-releasedevice, beads were arrayed and subsequently selected beads werereleased to form patterned lines. This entire procedure wasaccomplished within a few minutes. One characteristic of thisdevice revealed by the demonstration is that it is necessary torelease the beads in a predetermined order, from the upstreamto the downstream of the flow, during retrieval of multiple beadsfrom the array. In this case, beads were released sequentiallyfrom left to right. Failure to follow this order would result inreleased beads re-entering vacant traps farther downstream. Fig.3B shows the formation and dissipation of optical-based micro-bubbles during a typical run. During the application of the laser,the size of bubbles increased, and once the laser was switched off,these bubbles started to dissipate, shrinking in size. Removal ofheat by the flow, loss of heat through the thin cover glass, andthe use of porous polymer matrix [poly(dimethylsiloxane)] prob-ably contributed to the dissipation of the bubbles. After a bubblehas shrunk enough to pass through the narrow neck of the trap,it will usually be washed out by the flow. Bubbles that do not getwashed out will disappear after a few seconds. Also, as expected,higher flow rates lead to faster dissipation of the bubbles.Conventional microbubble-based devices (22) are beset withproblems; besides complicated fabrication process, wiring con-nections and control systems, residual bubbles can also causeobstructions if they do not dissipate after the heat source isswitched off. Difficulty in dissipation of these bubbles is attrib-uted to the formation of stable gas bubbles instead of vaporbubbles, due to the effervescence of dissolved gases from thesolution. In our device, bubbles formed in the traps and did notimpede the operation of the device in any way. These bubblesreadily dissipated and were removed during operation.

High-speed camera images captured the instant at which atrapped bead was displaced by an optical-based microbubble (Fig.3C, see SI Movie 3). IR laser set to a power of 0.3 W was focusedon the aluminum pattern (t � 0.0 s), and after 373 ms, bubbleformation started. The expanding bubble displaced the previouslyimmobilized bead into the main channel, where it was carried outof the array. This retrieval procedure typically took 0.6 s tocomplete. After the laser was switched off, the bubble cooled down,shrank and disappeared in �3 s. If the need arises, this emptied trapcan be used to immobilize another particle. Studies on the effect ofdifferent laser settings on bead release probability at different flowrates revealed that higher power settings at short intervals weremore efficient than low power settings at longer intervals (Fig. 3D).Higher release probability is attributed to the ease of bubbleformation because high power settings translate to higher heatgeneration and at short intervals, heat loss to forced convection islimited. Release efficiency can be further improved by reducing theflow rate. Also, in an automated system, it will be possible toimplement multiple retrieval attempts to increase the successprobability significantly.

Here, we will also demonstrate the potential of this device asa tool for screening methods such as the OBOC combinatoriallibrary method. In this demonstration, two kinds of beads,biotinylated and nonbiotinylated beads, were mixed and arrayed(Fig. 4A). These beads represented beads coated with differentchemical entities in the actual OBOC combinatorial librarymethod. When a solution of analyte, in this case, a solution ofstreptavidin conjugated with Alexa Fluor 546, was perfused overthe beads (10 nl/s for 3 min), streptavidin started to bind tobiotinylated beads, which could be seen as an increase influorescent response of these beads (Fig. 4B). These fluorescentbeads would correspond to positive beads in the actual OBOCcombinatorial library method. After 3 min of infusion, thestreptavidin flow was stopped and washing buffer was perfusedat 10 nl/s. We then selected a particular fluorescent (positive)bead and retrieved it from the array. Fig. 4C shows the image of

Fig. 2. Superimposed time-lapsed high speed camera images showing thepath taken by beads in devices A (A) and B (B). Arrows show the bypassing andtrapping modes.

Table 1. Geometric dimensions of �–Fluidic traps

W1, �m L1, �m W2, �m L2, �m H, �m Q1/Q2

Device A 6 5.5 25 163 28 0.76Device B 7.5 4.5 20 172.5 18 3.95

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f luorescent beads in the array taken 218 s after the infusion ofstreptavidin solution, and Fig. 4D shows the image taken aftera selected fluorescent bead was retrieved from the array. Thisbead could then be collected at the device’s outlet and sent forsubsequent structural analysis to identity the chemical com-pound on it. Existing OBOC assays in Petri dishes and methodsusing COPAS (4), an automated fluorescent activated bead-sorter, to screen OBOC combinatorial libraries are capable ofhandling large number of beads within a relatively short time.Compared with these methods, our microfluidic bead arraymethod offers a promising alternative because it only requiresminute amounts of reagents, and allows continual observation ofthe reaction throughout screening. Moreover, microfluidicsallows the possibility to generate a wide spectrum of concen-tration (26) from small volumes of reagents for screeningpurposes, which is challenging for other methods. Even forapplications that do not require post analyses such as chemicaldetection and clinical diagnostic purposes whereby identities ofthe probes are known in advance, our purposed device has itsadvantages. After the test, all of the beads can be removed andreplaced for another round of screening. Such a flexible andreusable format will greatly reduce the cost of bead-based assays.

Discussion will not be complete without considering thepossible denaturing of biomolecules caused by heating during theretrieval process. After the laser is switched off, the smallamount of heat stored in the aluminum heater is readily removedby an inflow of cooler water upstream (average velocity �7–14mm/s). This dissipation occurs over several hundred millisec-onds, which is the time scale for all of the water in the array tobe replaced during operation. Although the temperature of thealuminum heater is �132°C (22) during bubble formation, it isof no concern in cases where the microbubbles are merely usedto reset the device for reuse. As for screening methods such asthe OBOC combinatorial library method, where biomoleculescan be cleaved off the beads and analyzed, the condition of thesebiomolecules should have minimal impact on mass spectrometryanalysis. Moreover, only a small part of the bead will be actuallyin contact with the bubble, and for a very short span of time(300 ms) before it is brought away by the flow. Nevertheless,there are several ways to reduce the heat generated. The simplestway is to introduce cavities on the aluminum patterns to providefor controlled heterogeneous nucleation sites for bubble forma-tion (22). In addition, we can reduce the saturation temperature

Fig. 3. Bead retrieval experimental results. (A) Beads were selectively retrieved to form patterned lines to demonstrate the ease of operation of ourtrap-and-release device. In the first row, after every two beads, two consecutive beads were released to form a ‘‘dash line.’’ In the second row, after every fourbeads, two consecutive beads were released to form a ‘‘long-dash line.’’ Arrow indicates position of released beads. No beads were released from the third row,forming a ‘‘solid line’’ with the beads. All the beads were released from the fourth row to form an ‘‘empty line.’’ Flow direction of buffer is from left to right.Beads have to be released from the upstream to the downstream of the flow. In this case, the beads were released sequentially from left to right. The entireprocedure was accomplished within a few minutes using an automatic X–Y stage. (B) Graph showing dynamics of optical-based bubble during formation anddissipation. (C) Sequence of high speed camera images showing the retrieval mechanism using bubble. Photo showing trapped beads before application of laser(1), application of IR laser (2), bubble formation after 0.373 s (3), bubble displacing the bead out of the cavity (4), bead being carried along the loop channel (5),and bead moving toward the outlet (6). Bubble cooled down, shrank, and disappeared �3 s after the laser was switched off, and another bead could enter thetrap again. (D) Graph showing the success rate of bead release at different laser settings.

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of water in the device by switching from an infusion-based to awithdrawal-based (suction) system to drive fluids, resulting inthe lowering of temperature during bubble formation. Bothmethods will reduce the required laser power and applied timeinterval for bubble formation. Another approach would be to usethe expanding bubble to power a jet stream to eject the beadfrom the trap instead of using the bubble directly (22). Thisapproach creates a buffer zone between the bubble and the bead,insulating the bead from the heat produced.

ConclusionsOur main motivation was to develop robust and efficient meth-ods to manipulate particles and apply them to dynamic microar-ray applications. Our device consists of essentially two keystrategies: (i) hydrodynamic confinement based on the principleof fluidic resistance, and (ii) optical-based microbubbles. Theoptical approach does away with complicated on-chip integra-tion of circuits, greatly simplifying fabrication, packaging, andcontrol. The device is highly amenable to automatic processing,and can be easily scaled up (see SI Text) to cater for fast,high-throughput, and highly parallel screening. We envision sucha reliable, f lexible and reusable dynamic microarray platformthat allows transportation, immobilization, infusion of reagents,observation, and retrieval in a single integrated device will besought after by many in various fields. This tool if coupled withmethods such as OBOC combinatorial bead-library has thepotential to hasten research and discovery in the fields ofproteomics, diagnostics and drug discovery. With modifications(e.g., reduction in flow rate) to reduce mechanical stresses on

cells during immobilization, and use of biocompatible materials(e.g., gold or platinum) to replace the aluminum patches, webelieve that it will be possible to use this device as a platform forcell-based arrays (12, 13) to facilitate studies of pathological andphysiological phenomena in cells, which will have enormouspotential for cell-based diagnostic applications, drug testing andtoxicology studies. Although we presented both components inone device, they are by no means limited to be used jointly, norare they limited to microarray applications. For example, in thecase of disposable diagnostic chips, where retrieval of beads isnot necessary, an array made up of �-Fluidic traps alone will besufficient for observation purposes. As for the optical-basedmicrobubbles technique developed here, we believe it has manyother potential uses beyond microarrays applications; it can beused as actuators, valves, and pumps to replace conventionalbubbles formed by resistive heating (22, 23) or electrochemicalmeans (24).

Materials and MethodsDevice Fabrication. Aluminum heaters (�100 nm thick, 7.5 �m �15 �m) were patterned on cover glass (30 mm � 40 mm,thickness � 0.12–0.17 mm; Matsunami, Osaka, Japan) usingstandard lithography techniques, and the microchannels werefabricated with poly(dimethylsiloxane) (PDMS) (Sylgard 184;Dow Corning, Ithaca, NY) using soft lithography techniques.The aluminum patterns and PDMS slab were subsequentlyexposed to O2 plasma, aligned under microscope, and sealedirreversibly. We then baked the device on a hotplate for 1 h at76°C to strengthen the bonding. See SI Text for further details.

Fig. 4. Demonstration of the use of trap-and-release device in chemical microarray applications. (A) Biotinylated and nonbiotinylated beads were randomlyarrayed, and a solution of streptavidin conjugated with Alexa Fluor 546 was perfused over the beads. (B) Graph showing the increase in fluorescent responseof 10 of the beads (bead 91–100) in the device. White boxes in C show the observation windows (15 � 15 pixels) for these beads. The marked increase influorescence of the biotinylated beads can be easily distinguished from the nonbiotinylated beads (beads 92, 97, and 99). (C) Fluorescent image of the micro-beadarray taken at 218 s after the infusion of streptavidin solution. Arrow indicates the fluorescent bead to be retrieved. (D) Close-up fluorescent image of the device.Arrow indicates the position of the retrieved fluorescent bead (bead 98). (Scale bar, 100 �m.)

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Reagent Preparation. Twenty microliters of polystyrene beads(� � 15 �m, coefficient of variation � 2%; Fluka, Steinheim,Germany) were suspended in 1 ml of ultrapure water (specificresistance of 18 M�cm) with 4 �l of Tween 20 (Kanto Chemical,Tokyo, Japan) added as surfactant. This suspension was used forall of the experiments apart from the biotin-streptavidin dem-onstration. For this experiment, the suspension was prepared byadding 60 �l of biotinylated beads (� � 15.3 �m, coefficient ofvariation 4%, Sphero Biotin-polystyrene particles crosslinked,Spherotech, Libertyville, IL) and 5 �l of polystyrene beads(Fluka) to 500 �l of PBS solution with 1 �l of Tween 20. Thestreptavidin solution was prepared by dissolving StreptavidinAlexa Fluor 546 conjugate (Invitrogen, Carlsbad, CA) in PBS toobtain a concentration of 0.05 mg/ml PBS.

Device Operation. Micro manipulation system (Sigma Koki,Saitama, Japan) was used to create microbubbles in our exper-

iments. This system comprised an IR laser (1,064 nm), a lasercontroller for adjusting laser intensity, an automatic XY stagemanipulator, and a shutter controller for controlling the dura-tion of laser exposure. Syringe pumps (Micro 4; World PrecisionInstruments, Sarasota, FL) were used to control the flow rates,and reagents were infused into the device using 250- or 500-�lHamilton Gastight syringes (1700 series, TLL). Video imageswere taken with Hamamatsu CCD camera, and high-speedimages were captured with Photron high speed camera mountedonto an inverted microscope (Eclipse, TE300; Nikon, Tokyo).

We thank Dr. Yuichi Hiratsuka and Mr. Naoya Fukushima for helpfuldiscussions. W.-H.T. was supported by a Ministry of Education, Culture,Sports, Science, and Technology (MEXT), Government of Japan schol-arship. This work was supported in part by Grants-in-Aid for ScientificResearch on Priority Areas (innovative nanoscience) from MEXT,Japan.

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