Authors have no one to acknowledge.

1. Device Fabrication
<ol>
	<li>Design two photomasks, the first for patterning the surface acoustic wave (SAW) layer, and the second for the polydimethylsiloxane (PDMS) microchannel mold.
	<ol>
		<li>The first photomask has a pair of opposing interdigital transducers (IDTs)&#8212;also known as a SAW delay line&#8212;and markers for channel alignment and spatial reference during microscopy. In our standard device we have single-electrode IDTs with a finger width p = 10 &#956;m, aperture of 750 &#956;m, and 25 straight finger pairs. The resulting IDT generates SAWs with a wavelength &#955; = 4p = 40 &#956;m corresponding to an operating frequency fo = cSAW/&#955; &#8776; 100 MHz on 128&#176; YX lithium niobate (LN). Each IDT width should be above two times the width of the microchannel to reduce any misalignment effects while bonding the layers. IDT design parameters are discussed comprehensively in several books9-11. We remark that only one IDT (placed at the channel outlet) is necessary to drive the fluid into the channel in acoustic-counterflow, but patterning a full delay line assists in device testing.</li>
		<li>The second has a simple microchannel structure to be aligned along the SAW delay line, with a microchamber to form the channel inlet. In our typical devices, the channels have a width w = 300 mm and a length of 5 mm. As a general rule, the channel width should be at least 10&#955; to avoid diffraction effects during SAW propagation in the microchannel, however in our testing we found that a width of ~7&#955; would not significantly affect SAW propagation within the channel.</li>
	</ol>
	</li>
	<li>Begin with an LN wafer and cleave a 2 cm by 2 cm sample. In order to perform transmission microscopy it is necessary to use a double side polished wafer. Note that LN is a standard for its biocompatibility and the SAW polarization and high piezoelectric coupling coefficient along the major axis, however other piezoelectric materials may be used with appropriate design considerations.</li>
	<li>Clean the substrate by rinsing it in acetone, 2-propanol and drying with a nitrogen gun.</li>
	<li>Spin coat the sample with Shipley S1818 at 4,000 rpm for 1 min.</li>
	<li>Soft bake at 90 &#176;C for 1 min on a hot plate.</li>
	<li>Align the sample with the SAW layer mask using a mask aligner and expose it to UV light with a 55 mJ/cm2. Care should be taken to align the IDT direction along the major axis of the LN substrate.</li>
	<li>Rinse the sample in Microposit MF319 developer for 30 sec to remove the unexposed photoresist.</li>
	<li>Stop the development by rinsing the sample in deionized water and dry it with a nitrogen gun.</li>
	<li>Deposit a 10-nm-thick titanium adhesion layer followed by 100-nm-thick gold layer by thermal evaporation.</li>
	<li>Perform lift-off by sonicating the sample in acetone, then rinse it in 2-propanol and dry with a nitrogen gun.</li>
	<li>Silanize the device surface to make it hydrophobic in the microchannel area12.
	<ol>
		<li>Mask the microchamber area with AR-N-4340 negative tone photoresist by optical lithography according to the manufacturer&#39;s datasheet.</li>
		<li>Activate the sample surface with a 2 min oxygen plasma (Gambetti Kenologia Srl, Colibri) of 0.14 mbar pressure and 100 W power giving a bias voltage of approximately 450 V.</li>
		<li>Mix 35 ml hexadecane, 15 ml carbon tetrachloride (CCl4), and 20 &#956;l octadecyltrichlorosilane (OTS) into a beaker inside a fume hood. Place the device in the solution, and leave covered for two hours.</li>
		<li>Rinse the device with 2-propanol and dry it with a nitrogen gun.</li>
		<li>Check that the contact angle of water on the surface is above 90&#176;. If the contact angle is insufficient, clean the sample and re-perform the steps in 1.11.</li>
		<li>Remove the residual resist on the sample by rinsing in acetone, 2-propanol and drying with a nitrogen gun.</li>
	</ol>
	</li>
	<li>Mount the sample on a printed circuit board with radio frequency waveguides and standard coaxial connectors (RF-PCB), and then put acoustic absorber (First Contact polymer) on the sample edges and connect the IDT by wire bonding or using pogo connectors.</li>
	<li>A master mold of the channel layer is patterned with SU-8 onto a small piece of Silicon (Si) wafer using standard optical photolithography. SU-8 type and photolithography recipe will be dependent on the final PDMS internal channel height required.</li>
	<li>Cast PDMS on the mold
	<ol>
		<li>Mix PDMS with a curing agent at a ratio of 10:1.</li>
		<li>Centrifuge the PDMS for 2 min at 1,320 x g for degassing.</li>
		<li>Pour the PDMS gently onto the SU-8 mold in a Petri dish to a total PDMS height on the order of 1 mm. The open Petri dish can be placed in a vacuum desiccator for approximately 30 min in order to degas the PDMS further.</li>
		<li>Once degassed, cure PDMS by heating to 80 &#176;C for one hour in an oven. Note that baking time and temperature can affect the mechanical properties of PDMS.</li>
	</ol>
	</li>
	<li>Prepare the solid PDMS layer
	<ol>
		<li>Cut around the channel using a surgical blade, being careful not to damage the SU8 master, and peel it off.</li>
		<li>Replica edges are then refined and straightened using a razor blade leaving at least 2 mm clearance on lateral side of the channel and no clearance (cut right through) at the channel outlet.</li>
		<li>Punch a hole in the microchamber using a Harris Unicore puncher to form the fluid-loading inlet.</li>
	</ol>
	</li>
	<li>Bond the PDMS channel with the LN substrate by simple conformal bonding. In this way the bond will hold throughout the fluid testing stage while remaining reversible.
	<ol>
		<li>Both surfaces are cleaned prior to joining by blowing away any excess debris with compressed nitrogen air. It is critical when joining the pieces to align the channel with the major axis of the LN according to the patterned alignment marks.</li>
	</ol>
	</li>
	<li>The complete device schematic is shown in Figure 1. Store completed devices in a clean environment until use.</li>
</ol>
Note: It is important that all fabrication steps are carried out in a clean room environment to avoid contamination of the device before use.
Note: Any of the optical lithography steps may be replaced by the user preferred methods.
Note: The silanization procedure may be substituted for a preferred hydrophobic coating method13.
2. RF Device Testing
<ol>
	<li>Calibrate the network or spectrum analyzer with an open/short waveguide on your RF-PCB.</li>
	<li>Connect the SAW delay line to the ports of a spectrum analyzer and measure the scattering matrix of the device. The transmission for a pair of single-electrode transducers will resemble the absolute value of a sinc function centered at the operating frequency of the IDT. In the reflection spectrum a dip (minimum) is observed at the same frequency9-11. In our devices at 100 MHz operating frequency along the major axis typical values are -15 dB for S11 and S22 and -10 dB for S12 (without PDMS channels).</li>
</ol>
3. Microfluidics and Particle Flow Dynamics Visualization Experiment and Analysis
<ol>
	<li>Place the sample under a microscope. The specific optical setup depends on the SAW microfluidics phenomena to be observed. For example, a simple reflection microscope equipped with a 4X objective and a 30 fps video camera will be suitable to study fluid filling dynamics. To investigate more complex microparticle dynamics, it may be necessary to use a microscope equipped with a 20X objective and a 100 fps or higher video camera. It is important that both the objective and frame rate are high enough to capture any spatially and temporally important flow features.</li>
	<li>Connect the IDT in front of the channel outlet to an RF signal generator and operate it at the resonant frequency observed in the scattering matrix measurements. The typical operating power in acoustic-counterflow experiments is 20 dBm. If necessary, use a high-power UHF amplifier. Acoustic-streaming and atomization phenomena are observed without acoustic counterflow while running the device at lower power: typically acoustic-streaming recirculation begins at 0 dBm and atomization occurs above 14 dBm.</li>
	<li>Load 60 &#956;l of fluid into the microchamber with a micropipette. Fluid will passively diffuse into the microchamber. If necessary, gently push on the microchamber surface in order to favor the microchamber filling.
	<ol>
		<li>In order to visualize the flow it is necessary to add microbeads to the fluid. Note that in order to avoid particle clustering, sonicate the particle suspension prior to the experiments. To avoid particle adhesion on the substrate apply a 0 dBm signal to the device while loading.</li>
	</ol>
	</li>
	<li>Start recording the video through the microscope and increase the operating power in order to observe acoustic counterflow. Different flow schemes will be determined by input power, chip design and particle diameter.
	<ol>
		<li>In order to qualitatively capture the dynamics, fluid flow has to be recorded in proximity of the meniscus and inlet at different stages of channel filling using markers as a spatial reference.</li>
		<li>In order to perform quantitative measurement of particle dynamics by micro particle image velocimetry (&#956;PIV) 14,15 or spatial temporal image correlation spectroscopy (STICS) 16,17, fluid flow has to be recorded in the point of interest with a fixed field of view for at least 100 frames at a frame rate imposed by the particle dynamics.</li>
	</ol>
	</li>
	<li>Analyze the video with image processing software. The choice of the software to be used depends on the phenomena of interest. For example, to quantify the size distribution of atomized droplets, spatial periodicity of particle accumulation, or manual tracking of diluted particles, simple freeware image analysis software such as Fiji is suitable18; whereas in order to obtain streamlines and velocity field measurements, customized mPIV19 or STICS20 code is required. In our analysis customized STICS code is written in MATLAB, however a preferred alternative coding language may be equally acceptable.</li>
</ol>

<ol>
	<li>Masini, L., Cecchini, M., Girardo, S., Cingolani, R., Pisignano, D., Beltram, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface-acoustic-wave+counterflow+micropumps+for+on-chip+liquid+motion+control+in+two-dimensional+microchannel+arrays.">Surface-acoustic-wave counterflow micropumps for on-chip liquid motion control in two-dimensional microchannel arrays.</a> Lab on a Chip. 10 (15), 1997-2000 (2010).</li><li>Travagliati, M., De Simoni, G., Lazzarini, C. M., Piazza, V., Beltram, F., Cecchini, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interaction-free,+automatic,+on-chip+fluid+routing+by+surface+acoustic+waves.">Interaction-free, automatic, on-chip fluid routing by surface acoustic waves.</a> Lab on a Chip. 12 (15), 2621-2624 (2012).</li><li>Wixforth, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acoustically+driven+planar+microfluidics.">Acoustically driven planar microfluidics.</a> Superlattices and Microstructures. 33 (5), 389-396 (2003).</li><li>Friend, J., Yeo, L. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microscale+acoustofluidics:+Microfluidics+driven+via+acoustics+and+ultrasonics.">Microscale acoustofluidics: Microfluidics driven via acoustics and ultrasonics.</a> Reviews of Modern Physics. 83 (2), 647-64 (2011).</li><li>Tan, M. K., Yeo, L. Y., Friend, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+fluid+flow+and+mixing+induced+in+microchannels+using+surface+acoustic+waves.">Rapid fluid flow and mixing induced in microchannels using surface acoustic waves.</a> Europhysics Letters. 87, 47003 (2009).</li><li>Glass, N., Shilton, R., Chan, P., Friend, J., Yeo, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Miniaturised+Lab-on-a-Disc+(miniLOAD).">Miniaturised Lab-on-a-Disc (miniLOAD).</a> SMALL, Small. 8 (12), 1880-1880 (2012).</li><li>Madou, M. J., Kellogg, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=LabCD:+a+centrifuge-based+microfluidic+platform+for+diagnostics.">LabCD: a centrifuge-based microfluidic platform for diagnostics.</a> Proceedings of SPIE. 3259, 80 (1998).</li><li>Cecchini, M., Girardo, S., Pisignano, D., Cingolani, R., Beltram, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acoustic-counterflow+microfluidics+by+surface+acoustic+waves.">Acoustic-counterflow microfluidics by surface acoustic waves.</a> Applied Physics Letters. 92 (10), 104103 (2008).</li><li>Campbell, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Surface acoustic wave devices for mobile and wireless communications. 1, (1998).</li><li>Hashimoto, K. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Surface acoustic wave devices in telecommunications: modelling and simulation. , (2000).</li><li>Royer, D., Dieulesaint, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elastic+Waves+in+Solids+II.">Elastic Waves in Solids II.</a> Generation, Acousto-Optic Interaction, Applications. 2, (2000).</li><li>Renaudin, A., Sozanski, J. P., Verbeke, B., Zhang, V., Tabourier, P., Druon, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monitoring+SAW-actuated+microdroplets+in+view+of+biological+applications.">Monitoring SAW-actuated microdroplets in view of biological applications.</a> Sensors and Actuators B: Chemical. 138 (1), 374-382 (2009).</li><li>Glass, N. R., Tjeung, R., Chan, P., Yeo, L. Y., Friend, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Organosilane+deposition+for+microfluidic+applications.">Organosilane deposition for microfluidic applications.</a> Biomicrofluidics. 5 (3), 036501 (2011).</li><li>Wereley, S. T., Meinhart, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+advances+in+micro-particle+image+velocimetry.">Recent advances in micro-particle image velocimetry.</a> Annual Review of Fluid Mechanics. 42, 557-576 (2010).</li><li>Augustsson, P., Barnkob, R., Wereley, S. T., Bruus, H., Laurell, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+and+temperature-controlled+micro-PIV+measurements+enabling+long-term-stable+microchannel+acoustophoresis+characterization.">Automated and temperature-controlled micro-PIV measurements enabling long-term-stable microchannel acoustophoresis characterization.</a> Lab on a Chip. 11 (24), 4152-4164 (2011).</li><li>Hebert, B., Costantino, S., Wiseman, P. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatiotemporal+image+correlation+spectroscopy+(STICS)+theory,+verification,+and+application+to+protein+velocity+mapping+in+living+CHO+cells.">Spatiotemporal image correlation spectroscopy (STICS) theory, verification, and application to protein velocity mapping in living CHO cells.</a> Biophysical Journal. 88 (5), 3601 (2005).</li><li>Rossow, M., Mantulin, W. W., Gratton, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatiotemporal+image+correlation+spectroscopy+measurements+of+flow+demonstrated+in+microfluidic+channels.">Spatiotemporal image correlation spectroscopy measurements of flow demonstrated in microfluidic channels.</a> Journal of Biomedical Optics. 14 (2), 024014 (2009).</li><li>Schindelin, J., Arganda-Carreras, I., Frise, E., Kaynig, V., Longair, M., Pietzsch, T., Preibisch, S., Rueden, C., Saalfeld, S., Schmid, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fiji:+an+open-source+platform+for+biological-image+analysis.">Fiji: an open-source platform for biological-image analysis.</a> Nature Methods. 9 (7), 676-682 (2012).</li><li>Rogers, P. R., Friend, J. R., Yeo, L. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exploitation+of+surface+acoustic+waves+to+drive+size-dependent+microparticle+concentration+within+a+droplet.">Exploitation of surface acoustic waves to drive size-dependent microparticle concentration within a droplet.</a> Lab on a Chip. 10 (21), 2979-2985 (2010).</li><li>Muller, P. B., Barnkob, R., Jensen, M. J. H., Bruus, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+numerical+study+of+microparticle+acoustophoresis+driven+by+acoustic+radiation+forces+and+streaming-induced+drag+forces.">A numerical study of microparticle acoustophoresis driven by acoustic radiation forces and streaming-induced drag forces.</a> Lab on a Chip. 12 (22), 4617-4627 (2012).</li><li>Luo, J. K., Fu, Y. Q., Li, Y., Du, X. Y., Flewitt, A. J., Walton, A. J., Milne, W. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Moving-part-free+microfluidic+systems+for+lab-on-a-chip.">Moving-part-free microfluidic systems for lab-on-a-chip.</a> Journal of Micromechanics and Microengineering. 19 (5), 054001-05 (2009).</li><li>Lindken, R., Rossi, M., Gro&#223;e, S., Westerweel, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Micro-particle+image+velocimetry+(&#956;PIV):+recent+developments,+applications,+and+guidelines.">Micro-particle image velocimetry (&#956;PIV): recent developments, applications, and guidelines.</a> Lab on a Chip. 9 (17), 2551-2567 (2009).</li><li>Gedge, M., Hill, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acoustofluidics+17:+Theory+and+applications+of+surface+acoustic+wave+devices+for+particle+manipulation.">Acoustofluidics 17: Theory and applications of surface acoustic wave devices for particle manipulation.</a> Lab on a Chip. 12 (17), 2998-3007 (2012).</li></ol>

Authors have nothing to disclose.

One of the greatest challenges faced by the microfluidic community is the realization of an actuation platform for truly portable point-of-care devices. Among the proposed integrated micropumps23,those based on surface acoustic waves (SAWs) are particularly attractive due to their associated capabilities in fluid mixing, atomization and particle concentration and separation4. In this paper we have demonstrated how to fabricate and operate a lab-on-chip device in which fluid is steered in a closed PD...

One of the continued challenges facing the microfluidic community is the need to have an efficient pumping mechanism that can be miniaturized for integration into truly portable micro-total-analysis systems (&#956;TAS&#39;s). Standard macroscopic pumping systems simply fail to provide the portability required for &#956;TAS&#39;s, owing to the unfavorable scaling of the volumetric flow rates as the channel size decreases down to the micron range or below. On the contrary, SAWs have gained increasing interest as fluid actuation mechanisms and appear as a promising avenue for the solution of some of these problems1,2.
SAWs were shown to provide a very efficient mechanism of energy transport into fluids3. When a SAW propagates onto a piezoelectric substrate, e.g. lithium niobate (LN), the wave will be radiated into any fluid in its path at an angle known as the Rayleigh angle &#952;R = sin&#8722;1 (cf /cs), owing to the mismatch of sound velocities in the substrate, cs, and the fluid cf. This leakage of radiation into the fluid gives rise to a pressure wave which drives acoustic streaming in the fluid. Depending on the device geometry and power applied to the device, this mechanism was shown to actuate a wide variety of on-chip processes, such as fluid mixing, particle sorting, atomization, and pumping1,4. Despite the simplicity and effectiveness of actuating microfluids with SAW, there are only a small number of SAW driven microfluidic pumping mechanisms that have been demonstrated to date. The first demonstration was the simple translation of free droplets placed in the SAW propagation path on a piezoelectric substrate3. This novel method generated much interest in using SAWs as a microfluidic actuation method, however there was still a need for fluids to be driven through enclosed channels&#8212;a more difficult task. Tan et al. demonstrated pumping within a microchannel that was laser ablated directly into the piezoelectric substrate. By geometric modification with respect to the channel and IDT dimensions, they were able to demonstrate both uniform and mixing flows5. Glass et al. recently demonstrated a method of moving fluids through microchannels and microfluidic components by combining SAW actuated rotations with centrifugal microfluidics, as a demonstration of true miniaturization of the popular Lab-on-a-CD concept6,7. However, the only fully enclosed SAW driven pumping mechanism that has been demonstrated remains to be Cecchini et al.&#39;s SAW-driven acoustic counterflow8&#8212;the focus of this video. It exploits the atomization and coalescence of a fluid to pump it through a closed channel in the direction opposing the propagation direction of the acoustic wave. This system can give rise to surprisingly complex flows within a microchannel. Moreover, depending on the device geometry, it can provide a range of flow schemes, from laminar flows to more complex regimes characterized by vortices and particle-accumulation domains. The ability to easily influence the flow characteristics within the device shows opportunities for advanced on-chip particle manipulation.
In this protocol we wish to clarify the main aspects of practical SAW-based microfluidics: device fabrication, experimental operation, and flow visualization. While we are explicitly describing these procedures for the fabrication and operation of SAW-driven acoustic counterflow devices, these sections can easily be modified for their application to a range of SAW-driven microfluidic regimes.

<table border="1">
	<tbody>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Double side polished 128&#176; YX lithium niobate wafer</td>
			<td>Crystal Technology, LLC</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Silicon wafer</td>
			<td>Siegert Wafers</td>
			<td></td>
			<td>We use &#60;100&#62;</td>
		</tr>
		<tr>
			<td>IDT Optical lithography mask with alignment marks (positive)</td>
			<td>Any vendor</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Channel Optical lithography mask (negative)</td>
			<td>Any vendor</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Positive photoresist</td>
			<td>Shipley</td>
			<td>S1818</td>
			<td></td>
		</tr>
		<tr>
			<td>Positive photoresist developer</td>
			<td>Microposit</td>
			<td>MF319</td>
			<td></td>
		</tr>
		<tr>
			<td>Negative tone photoresist</td>
			<td>Allresist</td>
			<td>AR-N-4340</td>
			<td></td>
		</tr>
		<tr>
			<td>Negative tone photoresist developer</td>
			<td>Allresist</td>
			<td>AR 300-475</td>
			<td></td>
		</tr>
		<tr>
			<td>SU8 thick negative tone photoresist</td>
			<td>Microchem</td>
			<td>SU-8 2000 Series</td>
			<td></td>
		</tr>
		<tr>
			<td>SU8 thick negative tone photoresist developer</td>
			<td>Microchem</td>
			<td>SU-8 developer</td>
			<td></td>
		</tr>
		<tr>
			<td>Hexadecane</td>
			<td>Sigma-Aldrich</td>
			<td>H6703</td>
			<td></td>
		</tr>
		<tr>
			<td>Carbon tetrachloride (CCl4)</td>
			<td>Sigma-Aldrich</td>
			<td>107344</td>
			<td></td>
		</tr>
		<tr>
			<td>Octadecyltrichlorosilane (OTS)</td>
			<td>Sigma-Aldrich</td>
			<td>104817</td>
			<td></td>
		</tr>
		<tr>
			<td>Acetone CMOS grade</td>
			<td>Sigma-Aldrich</td>
			<td>40289</td>
			<td></td>
		</tr>
		<tr>
			<td>2-propanol CMOS grade</td>
			<td>Sigma-Aldrich</td>
			<td>40301</td>
			<td></td>
		</tr>
		<tr>
			<td>Titanium</td>
			<td>Any vendor</td>
			<td>99.9% purity</td>
			<td></td>
		</tr>
		<tr>
			<td>Gold</td>
			<td>Any vendor</td>
			<td>99.9% purity</td>
			<td></td>
		</tr>
		<tr>
			<td>PDMS</td>
			<td>Dow Corning</td>
			<td>Sylgard 184 silicone elastomer kit with curing agent</td>
			<td></td>
		</tr>
		<tr>
			<td>Petri dish</td>
			<td>Any vendor</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>5 mm ID Harris Uni-Core multi-purpose coring tool</td>
			<td>Sigma-Aldrich</td>
			<td>Z708895</td>
			<td>Any diameter greater than 2 mm is suitable</td>
		</tr>
		<tr>
			<td>Acoustic absorber</td>
			<td>Photonic Cleaning Technologies</td>
			<td>First Contact regular kit</td>
			<td></td>
		</tr>
		<tr>
			<td>RF-PCB</td>
			<td>Any vendor</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Spinner</td>
			<td>Laurell technologies corporation</td>
			<td>WS-400-6NPP</td>
			<td>Any spinner can be used</td>
		</tr>
		<tr>
			<td>UV Mask aligner</td>
			<td>Karl Suss</td>
			<td>MJB 4</td>
			<td>Any aligner can be used</td>
		</tr>
		<tr>
			<td>Thermal evaporator</td>
			<td>Kurt J. Lesker</td>
			<td>Nano 38</td>
			<td>Any thermal, e-beam evaporator or sputtering system can be used</td>
		</tr>
		<tr>
			<td>Oxygen plasma asher</td>
			<td>Gambetti Kenologia Srl</td>
			<td>Colibr&#236;</td>
			<td>Any plasma asher or RIE machine can be used</td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Eppendorf</td>
			<td>5810 R</td>
			<td>Any centrifuge can be used</td>
		</tr>
		<tr>
			<td>Wire bonder</td>
			<td>Kulicke &#38; Soffa</td>
			<td>4523AD</td>
			<td>Any wire bonder can be used if the PCB is used without pogo connectors</td>
		</tr>
		<tr>
			<td>Contact Angle Meter</td>
			<td>KSV</td>
			<td>CAM 101</td>
			<td>Any contact angle meter can be used</td>
		</tr>
		<tr>
			<td>Spectrum analyzer</td>
			<td>Anristu</td>
			<td>56100A</td>
			<td>Any spectrum or network analyzer can be used</td>
		</tr>
		<tr>
			<td>RF signal generator</td>
			<td>Anristu</td>
			<td>MG3694A</td>
			<td>Any RF signal generator can be used</td>
		</tr>
		<tr>
			<td>RF high power amplifier</td>
			<td>Mini Circuits</td>
			<td>ZHL-5W-1</td>
			<td>Any RF high power amplifier can be used</td>
		</tr>
		<tr>
			<td>Microbeads suspension</td>
			<td>Sigma-Aldrich</td>
			<td>L3280</td>
			<td>Depending on the experimental purpose different suspension of different diameter and different material properties can be used</td>
		</tr>
		<tr>
			<td>Optical microscope</td>
			<td>Nikon</td>
			<td>Ti-Eclipse</td>
			<td>Any optical microscope with spatial resolution satisfying experimental purposes can be used</td>
		</tr>
		<tr>
			<td>Video camera</td>
			<td>Basler</td>
			<td>A602-f</td>
			<td>Any video camera that has enough frame rate and sensitivity satisfying experimental purposes can be used</td>
		</tr>
		<tr>
			<td>Camera acquisition software</td>
			<td>Advanced technologies</td>
			<td>Motion Box</td>
			<td>Any software enabling high and controlled frame rate acquisition can be used</td>
		</tr>
	</tbody>
</table>

fabrication, operation and flow visualization in surface-acoustic-wave-driven acoustic-counterflow microfluidics

Surface acoustic waves (SAWs) can be used to drive liquids in portable microfluidic chips via the acoustic counterflow phenomenon. In this video we present the fabrication protocol for a multilayered SAW acoustic counterflow device. The device is fabricated starting from a lithium niobate (LN) substrate onto which two interdigital transducers (IDTs) and appropriate markers are patterned. A polydimethylsiloxane (PDMS) channel cast on an SU8 master mold is finally bonded on the patterned substrate. Following the fabrication procedure, we show the techniques that allow the characterization and operation of the acoustic counterflow device in order to pump fluids through the PDMS channel grid. We finally present the procedure to visualize liquid flow in the channels. The protocol is used to show on-chip fluid pumping under different flow regimes such as laminar flow and more complicated dynamics characterized by vortices and particle accumulation domains.

Figure 2 shows representative results of device RF testing which were taken prior to bonding the LN layer to the microchannel layer: typical S11 and S12 spectra are reported in panel a) and b) respectively. The depth of the valley at central frequency in S11 spectrum is related to the efficiency of conversion of RF power in SAW mechanical power. Hence, for a fixed number of IDT finger pairs, a reduction in the valley minimum will result i...

Watch this Scientific Journal Video about Fabrication, Operation and Flow Visualization in Surface-acoustic-wave-driven Acoustic-counterflow Microfluidics at JoVE.com

Fabrication, Operation and Flow Visualization in Surface-acoustic-wave-driven Acoustic-counterflow Microfluidics

In this video we first describe fabrication and operation procedures of a surface acoustic wave (SAW) acoustic counterflow device. We then demonstrate an experimental setup that allows for both qualitative flow visualization and quantitative analysis of complex flows within the SAW pumping device.

Surface acoustic waves (SAWs) can be used to drive liquids in portable microfluidic chips via the acoustic counterflow phenomenon. In this video we ...

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16.9K 조회수.  Istituto Italiano di Tecnologia. In this video we first describe fabrication and operation procedures of a surface acoustic wave (SAW) acoustic counterflow device. We then demonstrate an experimental setup that allows for both qualitative flow visualization and quantitative analysis of complex flows within the SAW pumping device.

비디오: Fabrication, Operation and Flow Visualization in Surface-acoustic-wave-driven Acoustic-counterflow Microfluidics

Synthesis and Operation of Fluorescent-core Microcavities for Refractometric Sensing

This paper discusses fluorescent core microcavity-based sensors that can operate in a microfluidic analysis setup. These structures are based on the formation of a fluorescent quantum-dot (QD) coating on the channel surface of a conventional microcapillary. Silicon QDs are especially attractive for this application, owing in part to their negligible toxicity compared to the II-VI and II-VI compound QDs, which are legislatively controlled substances in many countries. While the ensemble emission spectrum is broad and featureless, an Si-QD film on the channel wall of a capillary features a set of sharp, narrow peaks in the fluorescence spectrum, corresponding to the electromagnetic resonances for light trapped within the film. The peak wavelength of these resonances is sensitive to the external medium, thus permitting the device to function as a refractometric sensor in which the QDs never come into physical contact with the analyte. The experimental methods associated with the fabrication of the fluorescent-core microcapillaries are discussed in detail, as well as the analysis methods. Finally, a comparison is made between these structures and the more widely investigated liquid-core optical ring resonators, in terms of microfluidic sensing capabilities.

Fluorescent-core microcavity sensors employ a high-index quantum-dot coating in the channel of silica microcapillaries. Changes in the refractive index of fluids pumped into the capillary channel cause shifts in the microcavity fluorescence spectrum that can be used to analyze the channel medium.

Refractometric 감지 용 형광 코어 Microcavities의 합성 및 운영

This paper discusses  fluorescent core microcavity-based sensors that can operate in a microfluidic  analysis setup. These structures are based on the ...

연구

공학

Fabrication and Visualization of Capillary Bridges in Slit Pore Geometry

A procedure for creating and imaging capillary bridges in slit-pore geometry is presented. High aspect ratio hydrophobic pillars are fabricated and functionalized to render their top surfaces hydrophilic. The combination of a physical feature (the pillar) with a chemical boundary (the hydrophilic film on the top of the pillar) provides both a physical and chemical heterogeneity that pins the triple contact line, a necessary feature to create stable long but narrow capillary bridges. The substrates with the pillars are attached to glass slides and secured into custom holders. The holders are then mounted onto four axis microstages and positioned such that the pillars are parallel and facing each other. The capillary bridges are formed by introducing a fluid in the gap between the two substrates once the separation between the facing pillars has been reduced to a few hundred micrometers. The custom microstage is then employed to vary the height of the capillary bridge. A CCD camera is positioned to image either the length or the width of the capillary bridge to characterize the morphology of the fluid interface. Pillars with widths down to 250 &#181;m and lengths up to 70 mm were fabricated with this method, leading to capillary bridges with aspect ratios (length/width) of over 1001.

A procedure for creating and imaging capillary bridges in slit-pore geometry is presented. The creation of capillary bridges relies on the formation of pillars to provide a directional physical and chemical heterogeneity to pin the fluid. Capillary bridges are formed and manipulated using microstages and visualized using a CCD camera.

슬릿 모공 기하학에서 모세관 다리의 제작 및 시각화

A procedure for creating and imaging capillary bridges in slit-pore geometry is presented. High aspect ratio hydrophobic pillars are fabricated and ...

Taking Advantage of Reduced Droplet-surface Interaction to Optimize Transport of Bioanalytes in Digital Microfluidics

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of &#8220;lab-on-a-chip&#8221; platforms. Often, droplet motion relies on the wetting of a surface, directly associated with the application of an electric field; surface interactions, however, make motion dependent on droplet contents, limiting the breadth of applications of the technique.
Some alternatives have been presented to minimize this dependence. However, they rely on the addition of extra chemical species to the droplet or its surroundings, which could potentially interact with droplet moieties. Addressing this challenge, our group recently developed Field-DW devices to allow the transport of cells and proteins in DMF, without extra additives.
Here, the protocol for device fabrication and operation is provided, including the electronic interface for motion control. We also continue the studies with the devices, showing that multicellular, relatively large, model organisms can also be transported, arguably unaffected by the electric fields required for device operation.

The protocol for fabrication and operation of field dewetting devices (Field-DW) is described, as well as the preliminary studies of the effects of electric fields on droplet contents.

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of &#8220;lab-on-a-chip&#8221; ...

Fabrication and Operation of a Nano-Optical Conveyor Belt

The technique of using focused laser beams to trap and exert forces on small particles has enabled many pivotal discoveries in the nanoscale biological and physical sciences over the past few decades. The progress made in this field invites further study of even smaller systems and at a larger scale, with tools that could be distributed more easily and made more widely available. Unfortunately, the fundamental laws of diffraction limit the minimum size of the focal spot of a laser beam, which makes particles smaller than a half-wavelength in diameter hard to trap and generally prevents an operator from discriminating between particles which are closer together than one half-wavelength. This precludes the optical manipulation of many closely-spaced nanoparticles and&#160;limits the&#160;resolution of optical-mechanical systems. Furthermore, manipulation using focused beams requires beam-forming or steering optics, which can be very bulky and expensive. To address these limitations in the system scalability of conventional optical trapping our lab has devised an alternative technique which utilizes near-field optics to move particles across a chip. Instead of focusing laser beams in the far-field, the optical near field of plasmonic resonators produces the necessary local optical intensity enhancement to overcome the restrictions of diffraction and manipulate particles at higher resolution. Closely-spaced resonators produce strong optical traps which can be addressed to mediate the hand-off of particles from one to the next in a conveyor-belt-like fashion. Here, we describe how to design and produce a conveyor belt using a gold surface patterned with plasmonic C-shaped resonators and how to operate it with polarized laser light to achieve super-resolution nanoparticle manipulation and transport. The nano-optical conveyor belt chip can be produced using lithography techniques and easily packaged and distributed.

The scalability and resolution of conventional optical manipulation techniques are limited by diffraction. We circumvent the diffraction limit and describe a method of optically transporting nanoparticles across a chip using a gold surface patterned with a path of closely spaced C-shaped plasmonic resonators.

나노 광학 컨베이어 벨트의 제작 및 운영

The technique of using focused laser beams to trap and exert forces on small particles has enabled many pivotal discoveries in the nanoscale biological ...

Fabrication and Operation of Acoustofluidic Devices Supporting Bulk Acoustic Standing Waves for Sheathless Focusing of Particles

Acoustophoresis refers to the displacement of suspended objects in response to directional forces from sound energy. Given that the suspended objects must be smaller than the incident wavelength of sound and the width of the fluidic channels are typically tens to hundreds of micrometers across, acoustofluidic devices typically use ultrasonic waves generated from a piezoelectric transducer pulsating at high frequencies (in the megahertz range). At characteristic frequencies that depend on the geometry of the device, it is possible to induce the formation of standing waves that can focus particles along desired fluidic streamlines within a bulk flow. Here, we describe a method for the fabrication of acoustophoretic devices from common materials and clean room equipment. We show representative results for the focusing of particles with positive or negative acoustic contrast factors, which move towards the pressure nodes or antinodes of the standing waves, respectively. These devices offer enormous practical utility for precisely positioning large numbers of microscopic entities (e.g., cells) in stationary or flowing fluids for applications ranging from cytometry to assembly.

Acoustofluidic devices use ultrasonic waves within microfluidic channels to manipulate, concentrate and isolate suspended micro and nanoscopic entities. This protocol describes the fabrication and operation of such a device supporting bulk acoustic standing waves to focus particles in a central streamline without the aid of sheath fluids.

제조 및 Acoustofluidic 장치의 작동은 입자의 초점 쉬스에 대한 대량 음향 서 파도 지원

Acoustophoresis refers to the displacement of suspended objects in response to directional forces from sound energy. Given that the suspended objects must ...

Fabrication and Characterization of Superconducting Resonators

Superconducting microwave resonators are of interest for a wide range of applications, including for their use as microwave kinetic inductance detectors (MKIDs) for the detection of faint astrophysical signatures, as well as for quantum computing applications and materials characterization. In this paper, procedures are presented for the fabrication and characterization of thin-film superconducting microwave resonators. The fabrication methodology allows for the realization of superconducting transmission-line resonators with features on both sides of an atomically smooth single-crystal silicon dielectric. This work describes the procedure for the installation of resonator devices into a cryogenic microwave testbed and for cool-down below the superconducting transition temperature. The set-up of the cryogenic microwave testbed allows one to do careful measurements of the complex microwave transmission of these resonator devices, enabling the extraction of the properties of the superconducting lines and dielectric substrate (e.g., internal quality factors, loss and kinetic inductance fractions), which are important for device design and performance.

Superconducting microwave resonators are of interest for detection of light, quantum computing applications and materials characterization. This work presents a detailed procedure for fabrication and characterization of superconducting microwave resonator scattering parameters.

초전도 공진기의 제조 및 특성

Superconducting microwave resonators are of interest for a wide range of applications, including for their use as microwave kinetic inductance detectors ...

Fabrication of Polymer Microspheres for Optical Resonator and Laser Applications

This paper describes three methods of preparing fluorescent microspheres comprising &#960;-conjugated or non-conjugated polymers: vapor diffusion, interface precipitation, and mini-emulsion. In all methods, well-defined, micrometer-sized spheres are obtained from a self-assembling process in solution. The vapor diffusion method can result in spheres with the highest sphericity and surface smoothness, yet the types of the polymers able to form these spheres are limited. On the other hand, in the mini-emulsion method, microspheres can be made from various types of polymers, even from highly crystalline polymers with coplanar, &#960;-conjugated backbones. The photoluminescent (PL) properties from single isolated microspheres are unusual: the PL is confined inside the spheres, propagates at the circumference of the spheres via the total internal reflection at the polymer/air interface, and self-interferes to show sharp and periodic resonant PL lines. These resonating modes are so-called "whispering gallery modes" (WGMs). This work demonstrates how to measure WGM PL from single isolated spheres using the micro-photoluminescence (&#181;-PL) technique. In this technique, a focused laser beam irradiates a single microsphere, and the luminescence is detected by a spectrometer. A micromanipulation technique is then used to connect the microspheres one by one and to demonstrate the intersphere PL propagation and color conversion from coupled microspheres upon excitation at the perimeter of one sphere and detection of PL from the other microsphere. These techniques, &#181;-PL and micromanipulation, are useful for experiments on micro-optic application using polymer materials.

Protocols for the synthesis of microspheres from polymers, the manipulation of microspheres, and micro-photoluminescence measurements are presented.

광 공진기 및 레이저 응용을위한 고분자 미소 구의 제조

This paper describes three methods of preparing fluorescent microspheres comprising &#960;-conjugated or non-conjugated polymers: vapor diffusion, ...

Fabrication of Ultra-thin Color Films with Highly Absorbing Media Using Oblique Angle Deposition

Ultra-thin film structures have been studied extensively for use as optical coatings, but performance and fabrication challenges remain.&#160; We present an advanced method for fabricating ultra-thin color films with improved characteristics. The proposed process addresses several fabrication issues, including large area processing. Specifically, the protocol describes a process for fabricating ultra-thin color films using an electron beam evaporator for oblique angle deposition of germanium (Ge) and gold (Au) on silicon (Si) substrates.&#160; Film porosity produced by the oblique angle deposition induces color changes in the ultra-thin film. The degree of color change depends on factors such as deposition angle and film thickness. Fabricated samples of the ultra-thin color films showed improved color tunability and color purity. In addition, the measured reflectance of the fabricated samples was converted into chromatic values and analyzed in terms of color. Our ultra-thin film fabricating method is expected to be used for various ultra-thin film applications such as flexible color electrodes, thin film solar cells, and optical filters. Also, the process developed here for analyzing the color of the fabricated samples is broadly useful for studying various color structures.

We present a detailed method for fabricating ultra-thin color films with improved characteristics for optical coatings. The oblique angle deposition technique using an electron beam evaporator allows improved color tunability and purity. Fabricated films of Ge and Au on Si substrates were analyzed by reflectance measurements and color information conversion.

기울기 각도 증 착을 사용 하 여 미디어를 흡수 하는 높은 가진 매우 얇은 필름의 제조

Ultra-thin film structures have been studied extensively for use as optical coatings, but performance and fabrication challenges remain.&#160; We present ...

Flow-assisted Dielectrophoresis: A Low Cost Method for the Fabrication of High Performance Solution-processable Nanowire Devices

Flow-assisted dielectrophoresis (DEP) is an efficient self-assembly method for the controllable and reproducible positioning, alignment, and selection of nanowires. DEP is used for nanowire analysis, characterization, and for solution-based fabrication of semiconducting devices. The method works by applying an alternating electric field between metallic electrodes. The nanowire formulation is then dropped onto the electrodes which are on an inclined surface to create a flow of the formulation using gravity. The nanowires then align along the gradient of the electric field and in the direction of the liquid flow. The frequency of the field can be adjusted to select nanowires with superior conductivity and lower trap density. 
In this work, flow-assisted DEP is used to create nanowire field effect transistors. Flow-assisted DEP has several advantages: it allows selection of nanowire electrical properties; control of nanowire length; placement of nanowires in specific areas; control of orientation of nanowires; and control of nanowire density in the device. 
The technique can be expanded to many other applications such as gas sensors and microwave switches. The technique is efficient, quick, reproducible, and it uses a minimal amount of dilute solution making it ideal for the testing of novel nanomaterials. Wafer scale assembly of nanowire devices can also be achieved using this technique, allowing large numbers of samples for testing and large-area electronic applications.

In this paper, flow assisted dielectrophoresis is demonstrated for the self-assembly of nanowire devices. The fabrication of a silicon nanowire field effect transistor is shown as an example.

흐름 기반 Dielectrophoresis: 고성능 솔루션 처리 나노와이어 소자의 제조에 대 한 낮은 비용 방법

Flow-assisted dielectrophoresis (DEP) is an efficient self-assembly method for the controllable and reproducible positioning, alignment, and selection of ...

Experimental Investigation of the Flow Structure over a Delta Wing Via Flow Visualization Methods

It is well known that the flow field over a delta wing is dominated by a pair of counter rotating leading edge vortices (LEV). However, their mechanism is not well understood. The flow visualization technique is a promising non-intrusive method to illustrate the complex flow field spatially and temporally. A basic flow visualization setup consists of a high-powered laser and optic lenses to generate the laser sheet, a camera, a tracer particle generator, and a data processor. The wind tunnel setup, the specifications of devices involved, and the corresponding parameter settings are dependent on the flow features to be obtained.
Normal smoke wire flow visualization uses a smoke wire to demonstrate the flow streaklines. However, the performance of this method is limited by poor spatial resolution when it is conducted in a complex flow field. Therefore, an improved smoke flow visualization technique has been developed. This technique illustrates the large-scale global LEV flow field and the small-scale shear layer flow structure at the same time, providing a valuable reference for later detailed particle image velocimetry (PIV) measurement.
In this paper, the application of the improved smoke flow visualization and PIV measurement to study the unsteady flow phenomena over a delta wing is demonstrated. The procedure and cautions for conducting the experiment are listed, including wind tunnel setup, data acquisition, and data processing. The representative results show that these two flow visualization methods are effective techniques for investigating the three-dimensional flow field qualitatively and quantitatively.

Here, we present a protocol to observe unsteady vortical flows over a delta wing using a modified smoke flow visualization technique and investigate the mechanism responsible for the oscillations of the leading-edge vortex breakdown locations.

It is well known that the flow field over a delta wing is dominated by a pair of counter rotating leading edge vortices (LEV). However, their mechanism is ...