The authors are grateful to the University of California and the NANO3 facility at UC San Diego for provision of funds and facilities in support of this work. This work was performed in part at the San Diego Nanotechnology Infrastructure (SDNI) of UCSD, a member of the National Nanotechnology Coordinated Infrastructure, which is supported by the National Science Foundation (Grant ECCS&#8211;1542148). The work presented here was generously supported by a research grant from the W.M. Keck Foundation. The authors are also grateful for the support of this work by the Office of Naval Research (via Grant 12368098).

1. Nano-height channel mask preparation
<ol>
	<li>Photolithography: With a pattern describing the desired shape of the nanoheight channels (Figure 1B), use normal photolithography and lift-off procedures to produce nanoheight depressions in an LN wafer. These depressions will become nanoheight channels upon wafer bonding in a later step. 
	NOTE: The lateral dimensions of the nanoscale depressions are microscale in this protocol. Electron beam or He/Ne ion beam lithography can be used to fabricate channels with nanoscale lateral dimensions; Ga+-based ion beam lithography causes swelling and uneven substrate profiles23. The orientation of the two LN wafers should match, otherwise, thermal stress may cause the wafers or the bond between them to fail.</li>
	<li>Sputter deposition to protect regions from dry etching: Place the wafer into the sputter deposition system. Draw down the chamber vacuum to 5 x 10-6 mTorr, allow Ar to flow at 2.5 mTorr, and sputter Cr at 200 W to produce a 400-nm thick sacrificial mask where reactive ion etching will be prevented when used in step 3 below.</li>
	<li>Lift-off: Transfer the wafer into a beaker with sufficient acetone to completely immerse the wafer. Sonicate at medium intensity for 10 min. Rinse with DI water and dry the wafer with dry N2 flow.</li>
	<li>Dicing: Use a dicing saw to dice the entire wafer into individual chips with (typically) one nanoslit pattern per chip. 
	NOTE: The protocol can be paused here.</li>
</ol>
2. Nano-height channel fabrication
<ol>
	<li>Reactive ion etching (RIE): Use RIE to etch nanoscale depressions into the uncovered regions of the LN substrate. Regions left covered by sacrificial Cr will be protected from etching. Set the RIE power to 200W, heat the chamber to 50 &#176;C, draw down the chamber vacuum to 20 mTorr, set the SF6 flow rate to 10 sccm, and etch for 20 min to produce a 120-nm deep nanoslit in LN.</li>
	<li>Hole drilling for channel inlets and outlets: With double-sided tape, attach an etched LN chip to a small steel plate and the plate to the bottom of a petri dish. The Petri dish should be large enough to permit complete immersion of the LN chip and steel plate. Fill the Petri dish with water to fully immerse the chip. Attach a 0.5-mm diameter diamond drill bit to a drill press, and drill at a high speed of at least 10,000 rpm to machine the desired inlets and outlets. Drilling through a 0.5 mm thick substrate should take about 10 to 15 s24 (Figure 1B). 
	NOTE: Immersion while drilling prevents excessive local heating and particulate jamming at the drill site. Other types of drill bits are unlikely to work, and hand drilling is not possible at any speed to our knowledge. Drill bit rotation speeds of 10,000 rpm or greater are recommended to avoid shattering the LN.</li>
	<li>Cr wet etching: Use a diamond tip engraving pen to clearly mark the flat, unetched face of the drilled LN to keep track of which side the nanoheight channel is located in the remaining steps. Sonicate the chips in Cr etchant. 
	NOTE: The protocol can be paused here. It is exceedingly difficult to determine which side of the LN chip has the etched nanoscale depression after the Cr is removed. The sonication time depends upon the etching rate and the Cr mask thickness.</li>
</ol>
3. Room-temperature Plasma Activated Bonding
<ol>
	<li>Solvent cleaning LN chips: Collect chip pairs&#8212;one SAW device (fabricated by normal photolithography, sputter deposition, and lift-off procedures) and one etched nanoscale depression chip&#8212;together to prepare them for bonding. Immerse the chip pairs in a beaker of acetone placed in a sonication bath and sonicate for 2 min. Transfer the chips to methanol and sonicate for 1 min. Transfer the chips to DI water.</li>
	<li>Piranha cleaning: Prepare piranha acid in a glass beaker in a well-ventilated hood, dedicated to the use of acid, by adding H2O2 (30% in water) to H2SO4 (96%) at a ratio of 1:3. Place all chips in a Teflon holder. Place the holder in the beaker and immerse all chips into the piranha solution for 10 min, then rinse the chips and holder sequentially in two separate DI water baths. Dry the chips with dry N2 and immediately transfer them into oxygen (O2) plasma activation equipment, keeping them covered during handling to avoid contamination. 
	CAUTION: Piranha solutions are highly corrosive, are strongly oxidizing, and are dangerous. Follow the specific rules handling them at your institution, but at least take extreme care and wear the proper safety equipment. Upon completion of the work, the piranha solution must be cooled for at least one hour before pouring into a dedicated waste container. 
	NOTE: It is necessary to rinse the LN chips twice in two DI water baths. Rinsing them once leaves residue behind that will likely ruin the bonding. Gold electrodes are used for IDTs because of their good resistance to piranha solution.</li>
	<li>Plasma surface activation: Activate the chip surfaces using plasma with 120 W of power while exposed to O2 flow at 120 sccm for 150 s. Immediately transfer the samples to a fresh DI water bath for at least 2 min. 
	NOTE: Plasma surface treatment quickly followed by DI water immersion will form hydroxyl groups upon the LN surface, increasing its free surface energy to later promote bonding.</li>
	<li>Room-temperature bonding: Dry the samples with dry N2 flow and carefully lay the nanoslit chip onto the SAW device chip in the desired position. Realign to produce the desired orientation. Then use tweezers or similar to push down upon the sample from its center to initiate the bond. Gently push down in areas that failed to bond after the initial push. 
	NOTE: The bonding may be easily seen through the transparent LN. Bonded regions are entirely transparent. LN that is not double-side polished will be more difficult to assess.</li>
	<li>Heating after bonding: Place bonded samples in a sprung clamp to safely exert loads on it despite thermal expansion, and place the clamped samples into an oven at room temperature (25 &#176;C). Set the oven heating temperature to 300 &#176;C, ramp rate to 2 &#176;C/min maximum, dwell time to 2 h, and then to automatically shut off to allow it and the clamped samples within to naturally cool to room temperature. 
	NOTE: The protocol can be paused here. The bonding between hydroxyl groups produces water at the bond, and heating removes the water to drastically increase the bond strength. Modest clamping forces are sufficient. Attempting to bond two chips of different orientations or materials may cause cracks due to mismatched thermal expansion and consequent stress.</li>
</ol>
4. Experimental setup and testing
<ol>
	<li>Observation: Observe the nanoslit under an inverted microscope. Include and rotate a linear polarizing filter in the optical path to suitably block birefringence-based image doubling in the LN. Use ultrapure DI water via the inlet to observe fluid motion in the completed nanoslit. 
	NOTE: Ultrapure liquid is strongly recommended to prevent clogging, especially after evaporation.</li>
	<li>SAW actuation: Attach absorbers at the ends of the SAW device to prevent reflected acoustic waves. Use a signal generator to apply a sinusoidal electric field to the IDT at its resonance frequency of around 40 MHz. Use an amplifier to amplify the signal. Use an oscilloscope to measure the actual voltage, current, and power applied to the device. Record the fluid motion during SAW actuation within the nanoslit using a camera attached to the microscope.</li>
</ol>

<ol>
	<li>Eijkel, J. C., Van Den Berg, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanofluidics:+what+is+it+and+what+can+we+expect+from+it?.">Nanofluidics: what is it and what can we expect from it?.</a> Microfluidics and Nanofluidics. 1 (3), 249-267 (2005).</li><li>Longhurst, M. J., Quirke, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temperature-driven+pumping+of+fluid+through+single-walled+carbon+nanotubes.">Temperature-driven pumping of fluid through single-walled carbon nanotubes.</a> Nano Letters. 7 (11), 3324-3328 (2007).</li><li>Wang, B., Kr&#225;l, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coulombic+dragging+of+molecules+on+surfaces+induced+by+separately+flowing+liquids.">Coulombic dragging of molecules on surfaces induced by separately flowing liquids.</a> Journal of the American Chemical Society. 128 (50), 15984-15985 (2006).</li><li>Insepov, Z., Wolf, D., Hassanein, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanopumping+using+carbon+nanotubes.">Nanopumping using carbon nanotubes.</a> Nano Letters. 6 (9), 1893-1895 (2006).</li><li>Gong, X., 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=A+charge-driven+molecular+water+pump.">A charge-driven molecular water pump.</a> Nature Nanotechnology. 2 (11), 709 (2007).</li><li>Joseph, S., Aluru, N. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pumping+of+confined+water+in+carbon+nanotubes+by+rotation-translation+coupling.">Pumping of confined water in carbon nanotubes by rotation-translation coupling.</a> Physical Review Letters. 101 (6), 064502 (2008).</li><li>Rinne, K. F., Gekle, S., Bonthuis, D. J., Netz, R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoscale+pumping+of+water+by+AC+electric+fields.">Nanoscale pumping of water by AC electric fields.</a> Nano Letters. 12 (4), 1780-1783 (2012).</li><li>Eslamian, M., Saghir, M. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+thermophoretic+particle+separators:+numerical+analysis+and+simulation.">Novel thermophoretic particle separators: numerical analysis and simulation.</a> Applied Thermal Engineering. 59 (1-2), 527-534 (2013).</li><li>Miansari, M., 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=Acoustic+Nanofluidics+via+Room-Temperature+Lithium+Niobate+Bonding:+A+Platform+for+Actuation+and+Manipulation+of+Nanoconfined+Fluids+and+Particles.">Acoustic Nanofluidics via Room-Temperature Lithium Niobate Bonding: A Platform for Actuation and Manipulation of Nanoconfined Fluids and Particles.</a> Advanced Functional Materials. 26 (43), 7861-7872 (2016).</li><li>Minzioni, P., 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=Roadmap+for+optofluidics.">Roadmap for optofluidics.</a> Journal of Optics. 19 (9), 093003 (2017).</li><li>Connacher, W., 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=Micro/nano+acoustofluidics:+materials,+phenomena,+design,+devices,+and+applications.">Micro/nano acoustofluidics: materials, phenomena, design, devices, and applications.</a> Lab on a Chip. 18 (14), 1952-1996 (2018).</li><li>Ren, Z., 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=Etching+characteristics+of+LiNbO3+in+reactive+ion+etching+and+inductively+coupled+plasma.">Etching characteristics of LiNbO3 in reactive ion etching and inductively coupled plasma.</a> Journal of Applied Physics. 103 (3), 034109 (2008).</li><li>Winnall, S., Winderbaum, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lithium+niobate+reactive+ion+etching.">Lithium niobate reactive ion etching.</a> Defence Science and Technology Organization. , (2000).</li><li>Hu, H., Ricken, R., Sohler, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Etching+of+lithium+niobate:+micro-and+nanometer+structures+for+integrated+optics.">Etching of lithium niobate: micro-and nanometer structures for integrated optics.</a> Topical Meeting Photorefractive Materials, Effects, and Devices-Control of Light and Matter, Bad Honnef. , (2009).</li><li>Jackel, J. L., Howard, R. E., Hu, E. L., Lyman, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reactive+ion+etching+of+LiNbO3.">Reactive ion etching of LiNbO3.</a> Applied Physics Letters. 38 (11), 907-909 (1981).</li><li>Smith, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Investigation of nanoscale etching and poling of lithium niobate. , (2014).</li><li>Tomita, Y., Sugimoto, M., Eda, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+bonding+of+LiNbO3+single+crystals+for+optical+waveguides.">Direct bonding of LiNbO3 single crystals for optical waveguides.</a> Applied Physics Letters. 66 (12), 1484-1485 (1995).</li><li>Howlader, M. M. R., Suga, T., Kim, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Room+temperature+bonding+of+silicon+and+lithium+niobate.">Room temperature bonding of silicon and lithium niobate.</a> Applied Physics Letters. 89 (3), 031914 (2006).</li><li>Chang, C. M., 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=A+parametric+study+of+ICP-RIE+etching+on+a+lithium+niobate+substrate.">A parametric study of ICP-RIE etching on a lithium niobate substrate.</a> 10th IEEE International Conference on Nano/Micro Engineered and Molecular Systems. , 485-486 (2015).</li><li>Queste, S., 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=Deep+reactive+ion+etching+of+quartz,+lithium+niobate+and+lead+titanate.">Deep reactive ion etching of quartz, lithium niobate and lead titanate.</a> JNTE (Journ&#233;es Nationales sur les Technologies) Proceedings. , (2008).</li><li>Xu, J., Wang, C., Tian, Y., Wu, B., Wang, S., Zhang, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glass-on-LiNbO3+heterostructure+formed+via+a+two-step+plasma+activated+low-temperature+direct+bonding+method.">Glass-on-LiNbO3 heterostructure formed via a two-step plasma activated low-temperature direct bonding method.</a> Applied Surface Science. 459, 621-629 (2018).</li><li>Tulli, D., Janner, D., Pruneri, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Room+temperature+direct+bonding+of+LiNbO3+crystal+layers+and+its+application+to+high-voltage+optical+sensing.">Room temperature direct bonding of LiNbO3 crystal layers and its application to high-voltage optical sensing.</a> Journal of Micromechanics and Microengineering. 21 (8), 085025 (2011).</li><li>Sridhar, M., Maurya, D. K., 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=Focused+ion+beam+milling+of+microchannels+in+lithium+niobate.">Focused ion beam milling of microchannels in lithium niobate.</a> Biomicrofluidics. 6 (012819), (2012).</li></ol>

The authors have nothing to disclose.

Room-temperature bonding is key to fabricating SAW-integrated nanoslit devices. Five aspects need to be considered to ensure successful bonding and sufficient bonding strength.
Time and power for plasma surface activation 
Increasing the plasma power will help increase the surface energy and accordingly increase the bonding strength. But the downside of increasing the power during plasma surface activation is the increase in surface roughness, which may adversely affect t...

Controllable nanoscale fluid transport in nanochannels&#8212;nanofluidics1&#8212;occurs on the same length scales as most biological macromolecules, and is promising for biological analysis and sensing, medical diagnosis, and material processing. Various designs and simulations have been developed in nanofluidics to manipulate fluids and particle suspensions based on temperature gradients2, Coulomb dragging3, surface waves4, static electric fields5,6,7, and thermophoresis8 over the last fifteen years. Recently, SAW has been shown9 to produce nanoscale fluid pumping and draining with sufficient acoustic pressure to overcome the dominance of surface and viscous forces that otherwise prevent effective fluid transport in nanochannels. The key benefit of acoustic streaming is its ability to drive useful flow in nanostructures without concern over the details of the chemistry of the fluid or particle suspension, making devices that utilize this technique immediately useful in biological analysis, sensing, and other physicochemical applications.
Fabrication of SAW-integrated nanofluidic devices requires fabrication of the electrodes&#8212;the interdigital transducer (IDT)&#8212;on a piezoelectric substrate, lithium niobate10, to facilitate generating the SAW. Reactive ion etching (RIE) is used to form a nanoscale depression in a separate LN piece, and LN-LN bonding of the two pieces produces a useful nanochannel. The fabrication process for SAW devices has been presented in many publications, whether using normal or lift-off ultraviolet photolithography alongside metal sputter or evaporation deposition11. For the LN RIE process to etch a channel in a specific shape, the effects on the etch rate and the channel&#39;s final surface roughness from choosing different LN orientations, mask materials, gas flow, and plasma power have been investigated12,13,14,15,16. Plasma surface activation has been used to significantly increase surface energy and hence improve the strength of bonding in oxides such as LN17,18,19,20. It is likewise possible to heterogeneously bond LN with other oxides, such as SiO2 (glass) via a two-step plasma activated bonding method21. Room-temperature LN-LN bonding, in particular, has been investigated using different cleaning and surface activation treatments22.
Here, we describe in detail the process to fabricate 40 MHz SAW-integrated 100-nm height nanochannels, often called nanoslit channels (Figure 1A). Effective fluid capillary filling and fluid draining by SAW actuation demonstrates the validity of both nanoslit fabrication and SAW performance in such a nanoscale channel. Our approach offers a nano-acoustofluidic system enabling investigation of a variety of physical problems and biological applications.

<table><tbody><tr><td>Absorber</td><td>Dragon Skin, Smooth-On, Inc., Macungie, PA, USA</td><td>Dragon Skin 10 MEDIUM</td><td></td></tr><tr><td>Amplifier</td><td>Mini-Circuits, Brooklyn, NY, USA</td><td>ZHL&amp;ndash;1&amp;ndash;2W&amp;ndash;S+</td><td></td></tr><tr><td>Camera</td><td>Nikon, Minato, Tokyo, Japan</td><td>D5300</td><td></td></tr><tr><td>Developer</td><td>Futurrex, NJ, USA</td><td>RD6</td><td></td></tr><tr><td>Diamond tip engraving pen</td><td>Malco, Memphis, TN, USA</td><td>Malco A50 USA Made Carbide Tipped Scribe</td><td></td></tr><tr><td>Dicing saw</td><td>Disco, Tokyo, Japan</td><td>Disco Automatic Dicing Saw 3220</td><td></td></tr><tr><td>Heating oven</td><td>Carbolite, Hope Valley, UK</td><td>HTCR 6/28</td><td>High Temperature Clean Room Oven - HTCR</td></tr><tr><td>Hole driller</td><td>Dremel, Mount Prospect, Illinois</td><td>Model #4000</td><td>4000 High Performance Variable Speed Rotary</td></tr><tr><td>Inverted microscope</td><td>Amscope, Irvine, CA, USA</td><td>IN480TC-FL-MF603</td><td></td></tr><tr><td>Lithium niobate substrate</td><td>PMOptics, Burlington, MA, USA</td><td>PWLN-431232</td><td>4&quot; double-side polished 0.5 mm thick 128&amp;deg; Y-rotated cut lithium niobate</td></tr><tr><td>Mask aligner</td><td>Heidelberg Instruments, Heidelberg, Germany</td><td>MLA150</td><td></td></tr><tr><td>Nano3 cleanroom facility</td><td>UCSD, La Jolla, CA, USA</td><td></td><td>Fabrication process is performed in it.</td></tr><tr><td>Negative photoresist</td><td>Futurrex, NJ, USA</td><td>NR9-1500PY</td><td></td></tr><tr><td>Oscilloscope</td><td>Keysight Technologies, Santa Rosa, CA, USA</td><td>InfiniiVision 2000 X-Series</td><td></td></tr><tr><td>Plasma surface activation</td><td>PVA TePla, Corona, CA, USA</td><td>PS100</td><td>Tepla Asher</td></tr><tr><td>Polarizer sheet</td><td>Edmund Optics, Barrington, NJ, USA</td><td>#86-182</td><td></td></tr><tr><td>RIE etcher</td><td>Oxford Instruments, Abingdon, UK</td><td>Plasmalab 100</td><td></td></tr><tr><td>Signal generator</td><td>NF Corporation, Yokohama, Japan</td><td>WF1967 multifunction generator</td><td></td></tr><tr><td>Sputter deposition</td><td>Denton Vacuum, NJ, USA</td><td>Denton 18</td><td>Denton Discovery 18 Sputter System</td></tr><tr><td>Teflon wafer dipper</td><td>ShapeMaster, Ogden, IL, USA</td><td>SM4WD1</td><td>Wafer Dipper 4&quot;</td></tr></tbody></table>

fabrication of nanoheight channels incorporating surface acoustic wave actuation via lithium niobate for acoustic nanofluidics

Controlled nanoscale manipulation of fluids is known to be exceptionally difficult due to the dominance of surface and viscous forces. Megahertz-order surface acoustic wave (SAW) devices generate tremendous acceleration on their surface, up to 108 m/s2, in turn responsible for many of the observed effects that have come to define acoustofluidics: acoustic streaming and acoustic radiation forces. These effects have been used for particle, cell, and fluid manipulation at the microscale, although more recently SAW has been used to produce similar phenomena at the nanoscale through an entirely different set of mechanisms. Controllable nanoscale fluid manipulation offers a broad range of opportunities in ultrafast fluid pumping and biomacromolecule dynamics useful for physical and biological applications. Here, we demonstrate nanoscale-height channel fabrication via room-temperature lithium niobate (LN) bonding integrated with a SAW device. We describe the entire experimental process including nano-height channel fabrication via dry etching, plasma-activated bonding on lithium niobate, the appropriate optical setup for subsequent imaging, and SAW actuation. We show representative results for fluid capillary filling and fluid draining in a nanoscale channel induced by SAW. This procedure offers a practical protocol for nanoscale channel fabrication and integration with SAW devices useful to build upon for future nanofluidics applications.

We perform fluid capillary filing and SAW-induced fluid draining in nano-height LN slits after successful fabrication and bonding of SAW integrated nanofluidic devices. Surface acoustic waves are generated by IDTs actuated by an amplified sinusoidal signal at the IDTs&#39; resonance frequency of 40 MHz, and the SAW propagates into the nanoslit via a piezoelectric LN substrate. The behavior of the fluid in the nanoslit interacting with SAW may be observed using an inverted microscope.
We demons...

Watch this Scientific Journal Video about Fabrication of Nanoheight Channels Incorporating Surface Acoustic Wave Actuation via Lithium Niobate for Acoustic Nanofluidics at JoVE.com

Fabrication of Nanoheight Channels Incorporating Surface Acoustic Wave Actuation via Lithium Niobate for Acoustic Nanofluidics

We demonstrate fabrication of nanoheight channels with the integration of surface acoustic wave actuation devices upon lithium niobate for acoustic nanofluidics via liftoff photolithography, nano-depth reactive ion etching, and room-temperature plasma surface-activated multilayer bonding of single-crystal lithium niobate, a process similarly useful for bonding lithium niobate to oxides.

Controlled nanoscale manipulation of fluids is known to be exceptionally difficult due to the dominance of surface and viscous forces. Megahertz-order ...

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5.7K Views.  University of California San Diego. Ons protocol biedt een gedetailleerde fabricagemethode van nanohoogtekanalen met oppervlakteakoestische golfactuatie via lithium niobate voor akoestische nanofluidics. Deze techniek kan worden gebruikt om ruimtetemperatuur plasma oppervlak geactiveerd meerlaagse hechting van single crystal lithium niobate uit te voeren, een proces dat even nuttig is voor het plakken van lithium niobate of siliciumdioxide en andere oxiden. Vuil en deeltjes moeten tijdens de reinigings- en plasmaoppervlaktegeac...

Video: Fabrication of Nanoheight Channels Incorporating Surface Acoustic Wave Actuation via Lithium Niobate for Acoustic Nanofluidics

Construction and Testing of Coin Cells of Lithium Ion Batteries

Rechargeable lithium ion batteries have wide applications in electronics, where customers always demand more capacity and longer lifetime. Lithium ion batteries have also been considered to be used in electric and hybrid vehicles1 or even electrical grid stabilization systems2. All these applications simulate a dramatic increase in the research and development of battery materials3-7, including new materials3,8, doping9, nanostructuring10-13, coatings or surface modifications14-17 and novel binders18. Consequently, an increasing number of physicists, chemists and materials scientists have recently ventured into this area. Coin cells are widely used in research laboratories to test new battery materials; even for the research and development that target large-scale and high-power applications, small coin cells are often used to test the capacities and rate capabilities of new materials in the initial stage. 
 In 2010, we started a National Science Foundation (NSF) sponsored research project to investigate the surface adsorption and disordering in battery materials (grant no. DMR-1006515). In the initial stage of this project, we have struggled to learn the techniques of assembling and testing coin cells, which cannot be achieved without numerous help of other researchers in other universities (through frequent calls, email exchanges and two site visits). Thus, we feel that it is beneficial to document, by both text and video, a protocol of assembling and testing a coin cell, which will help other new researchers in this field. This effort represents the "Broader Impact" activities of our NSF project, and it will also help to educate and inspire students.
In this video article, we document a protocol to assemble a CR2032 coin cell with a LiCoO2 working electrode, a Li counter electrode, and (the mostly commonly used) polyvinylidene fluoride (PVDF) binder. To ensure new learners to readily repeat the protocol, we keep the protocol as specific and explicit as we can. However, it is important to note that in specific research and development work, many parameters adopted here can be varied. First, one can make coin cells of different sizes and test the working electrode against a counter electrode other than Li. Second, the amounts of C black and binder added into the working electrodes are often varied to suit the particular purpose of research; for example, large amounts of C black or even inert powder were added to the working electrode to test the "intrinsic" performance of cathode materials14. Third, better binders (other than PVDF) have also developed and used18. Finally, other types of electrolytes (instead of LiPF6) can also be used; in fact, certain high-voltage electrode materials will require the uses of special electrolytes7.

A protocol to construct and test coin cells of lithium ion batteries is described. The specific procedures of making a working electrode, preparing a counter electrode, assembling a cell inside a glovebox and testing the cell are presented.

Rechargeable lithium ion batteries have wide applications in electronics, where customers always demand more capacity and longer lifetime.  Lithium ion ...

Synthesis of Phase-shift Nanoemulsions with Narrow Size Distributions for Acoustic Droplet Vaporization and Bubble-enhanced Ultrasound-mediated Ablation

High-intensity focused ultrasound (HIFU) is used clinically to thermally ablate tumors. To enhance localized heating and improve thermal ablation in tumors, lipid-coated perfluorocarbon droplets have been developed which can be vaporized by HIFU. The vasculature in many tumors is abnormally leaky due to their rapid growth, and nanoparticles are able to penetrate the fenestrations and passively accumulate within tumors. Thus, controlling the size of the droplets can result in better accumulation within tumors. In this report, the preparation of stable droplets in a phase-shift nanoemulsion (PSNE) with a narrow size distribution is described. PSNE were synthesized by sonicating a lipid solution in the presence of liquid perfluorocarbon. A narrow size distribution was obtained by extruding the PSNE multiple times using filters with pore sizes of 100 or 200 nm. The size distribution was measured over a 7-day period using dynamic light scattering. Polyacrylamide hydrogels containing PSNE were prepared for in vitro experiments. PSNE droplets in the hydrogels were vaporized with ultrasound and the resulting bubbles enhanced localized heating. Vaporized PSNE enables more rapid heating and also reduces the ultrasound intensity needed for thermal ablation. Thus, PSNE is expected to enhance thermal ablation in tumors, potentially improving therapeutic outcomes of HIFU-mediated thermal ablation treatments.

Phase-shift nanoemulsions (PSNE) can be vaporized using high intensity focused ultrasound to enhance localized heating and improve thermal ablation in tumors. In this report, the preparation of stable PSNE with a narrow size distribution is described. Furthermore, the impact of vaporized PSNE on ultrasound-mediated ablation is demonstrated in tissue-mimicking phantoms.

High-intensity  focused ultrasound (HIFU) is used clinically to thermally ablate tumors. To  enhance localized heating and improve thermal ablation in ...

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.

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 ...

Fabrication of Uniform Nanoscale Cavities via Silicon Direct Wafer Bonding

Measurements of the heat capacity and superfluid fraction of confined 4He have been performed near the lambda transition using lithographically patterned and bonded silicon wafers. Unlike confinements in porous materials often used for these types of experiments3, bonded wafers provide predesigned uniform spaces for confinement. The geometry of each cell is well known, which removes a large source of ambiguity in the interpretation of data.
Exceptionally flat, 5 cm diameter, 375 &#181;m thick Si wafers with about 1 &#181;m variation over the entire wafer can be obtained commercially (from Semiconductor Processing Company, for example). Thermal oxide is grown on the wafers to define the confinement dimension in the z-direction. A pattern is then etched in the oxide using lithographic techniques so as to create a desired enclosure upon bonding. A hole is drilled in one of the wafers (the top) to allow for the introduction of the liquid to be measured. The wafers are cleaned2 in RCA solutions and then put in a microclean chamber where they are rinsed with deionized water4. The wafers are bonded at RT and then annealed at ~1,100 &#176;C. This forms a strong and permanent bond. This process can be used to make uniform enclosures for measuring thermal and hydrodynamic properties of confined liquids from the nanometer to the micrometer scale.

A method for permanently bonding two silicon wafers so as to realize a uniform enclosure is described. This includes wafer preparation, cleaning, RT bonding, and annealing processes. The resulting bonded wafers (cells) have uniformity of enclosure ~1%1,2. The resulting geometry allows for measurements of confined liquids and gasses.

Measurements of the heat capacity and superfluid fraction of confined 4He have been performed near the lambda transition using lithographically patterned ...

Patterning via Optical Saturable Transitions - Fabrication and Characterization

This protocol describes the fabrication and characterization of nanostructures using a novel nanolithographic technique called Patterning via Optical Saturable Transitions (POST). In this technique the chemical properties of organic photochromic molecules that undergo single-photon reactions are exploited, enabling rapid top-down nanopatterning over large areas at low light intensities, thereby, allowing for the circumvention of the far-field diffraction barrier.4 Simple, cost-effective, high throughput and resolution alternatives to nanopatterning are being explored, such as, two-photon polymerization5,6, beam pen lithography (BPL)7, scanning electron beam lithography (SEBL), and focused ion beam (FIB) patterning. However, multi-photon approaches require high light intensities, which limit their potential for high throughput and offer low image contrast. Although, electron and ion beam lithographic processes offer increased resolution, the serial nature of the process is limited to slow writing speeds, which also prevents patterning of features over large areas. Beam-pen lithography is an approach towards parallel near-field optical lithography. However, the gap between the source of the beam and the surface of the photoresist needs to be controlled extremely precisely for good pattern uniformity and this is very challenging to accomplish for large arrays of beams. Patterning via Optical Saturable Transitions (POST) is an alternative optical nanopatterning technique for patterning sub-wavelength features1-3. Since this technique uses single photons instead of electrons, it is extremely fast and does not require high light intensities1-3, opening the door to massive parallelization.

We report that the diffraction limit of conventional optical lithography can be overcome by exploiting the transitions of organic photochromic derivatives induced by their photoisomerization at low light intensities.1-3 This paper outlines our fabrication technique and two locking mechanisms, namely: dissolution of one photoisomer and electrochemical oxidation.

This protocol describes the fabrication and characterization of nanostructures using a novel nanolithographic technique called Patterning via Optical ...

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.

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

Microparticle Manipulation by Standing Surface Acoustic Waves with Dual-frequency Excitations

We demonstrate a method for increasing the tuning ability of a standing surface acoustic wave (SSAW) for microparticles manipulation in a lab-on-a-chip (LOC) system. The simultaneous excitation of the fundamental frequency and its third harmonic, which is termed as dual-frequency excitation, to a pair of interdigital transducers (IDTs) could generate a new type of standing acoustic waves in a microfluidic channel. Varying the power and the phase in the dual-frequency excitation signals results in a reconfigurable field of the acoustic radiation force applied to the microparticles across the microchannel (e.g., the number and location of the pressure nodes and the microparticle concentrations at the corresponding pressure nodes). This article demonstrates that the motion time of the microparticle to only one pressure node can be reduced ~2-fold at the power ratio of the fundamental frequency greater than ~90%. In contrast, there are three pressure nodes in the microchannel if less than this threshold. Furthermore, adjusting the initial phase between the fundamental frequency and the third harmonic results in different motion rates of the three SSAW pressure nodes, as well as in the percentage of microparticles at each pressure node in the microchannel. There is a good agreement between the experimental observation and the numerical predictions. This novel excitation method can easily and non-invasively integrate into the LOC system, with a wide tenability and only a few changes to the experimental set-up.

A protocol for manipulating the microparticles in a microfluidic channel with a dual-frequency excitation is presented.

We demonstrate a method for increasing the tuning ability of a standing surface acoustic wave (SSAW) for microparticles manipulation in a lab-on-a-chip ...

A Microfluidic Platform for Precision Small-volume Sample Processing and Its Use to Size Separate Biological Particles with an Acoustic Microdevice

A major advantage of microfluidic devices is the ability to manipulate small sample volumes, thus reducing reagent waste and preserving precious sample. However, to achieve robust sample manipulation it is necessary to address device integration with the macroscale environment. To realize repeatable, sensitive particle separation with microfluidic devices, this protocol presents a complete automated and integrated microfluidic platform that enables precise processing of 0.15&#8211;1.5 ml samples using microfluidic devices. Important aspects of this system include modular device layout and robust fixtures resulting in reliable and flexible world to chip connections, and fully-automated fluid handling which accomplishes closed-loop sample collection, system cleaning and priming steps to ensure repeatable operation. Different microfluidic devices can be used interchangeably with this architecture. Here we incorporate an acoustofluidic device, detail its characterization, performance optimization, and demonstrate its use for size-separation of biological samples. By using real-time feedback during separation experiments, sample collection is optimized to conserve and concentrate sample. Although requiring the integration of multiple pieces of equipment, advantages of this architecture include the ability to process unknown samples with no additional system optimization, ease of device replacement, and precise, robust sample processing.

This protocol describes a system architecture for performing automated small volume (0.15&#8211;1.5 ml) particle separations using a microfluidic device, and discusses methods to optimize acoustofluidic device performance and operation.

A major advantage of microfluidic devices is the ability to manipulate small sample volumes, thus reducing reagent waste and preserving precious sample. ...

Bioengineering

Fabrication of Surface Acoustic Wave Devices on Lithium Niobate

Manipulation of fluids and particles by acoustic actuation at small scale is aiding the rapid growth of lab-on-a-chip applications. Megahertz-order surface acoustic wave (SAW) devices generate enormous accelerations on their surface, up to 108 m/s2, in turn responsible for many of the observed effects that have come to define acoustofluidics: acoustic streaming and acoustic radiation forces. These effects have been used for particle, cell, and fluid handling at the microscale&#8212;and even at the nanoscale. In this paper we explicitly demonstrate two major fabrication methods of SAW devices on lithium niobate: the details of lift-off and wet etching techniques are described step-by-step. Representative results for the electrode pattern deposited on the substrate as well as the performance of SAW generated on the surface are displayed in detail. Fabrication tricks and troubleshooting are covered as well. This procedure offers a practical protocol for high frequency SAW device fabrication and integration for future microfluidics applications.

Two fabrication techniques, lift-off and wet etching, are described in producing interdigital electrode transducers upon a piezoelectric substrate, lithium niobate, widely used to generate surface acoustic waves now finding broad utility in micro to nanoscale fluidics. The as-produced electrodes are shown to efficiently induce megahertz order Rayleigh surface acoustic waves.&#160;

Manipulation of fluids and particles by acoustic actuation at small scale is aiding the rapid growth of lab-on-a-chip applications. Megahertz-order ...

Fabrication and Characterization of Thickness Mode Piezoelectric Devices for Atomization and Acoustofluidics

We present a technique to fabricate simple thickness mode piezoelectric devices using lithium niobate (LN). Such devices have been shown to atomize liquid more efficiently, in terms of &#64258;ow rate per power input, than those that rely on Rayleigh waves and other modes of vibration in LN or lead zirconate titanate (PZT). The complete device is composed of a transducer, a transducer holder, and a &#64258;uid supply system. The fundamentals of acoustic liquid atomization are not well known, so techniques to characterize the devices and to study the phenomena are also described. Laser Doppler vibrometry (LDV) provides vibration information essential in comparing acoustic transducers and, in this case, indicates whether a device will perform well in thickness vibration. It can also be used to &#64257;nd the resonance frequency of the device, though this information is obtained more quickly via impedance analysis. Continuous &#64258;uid atomization, as an example application, requires careful &#64258;uid flow control, and we present such a method with high-speed imaging and droplet size distribution measurements via laser scattering.

Fabrication of piezoelectric thickness mode transducers via direct current sputtering of plate electrodes on lithium niobate is described. Additionally, reliable operation is achieved with a transducer holder and &#64258;uid supply system and characterization is demonstrated via impedance analysis, laser doppler vibrometry, high-speed imaging, and droplet size distribution using laser scattering.

We present a technique to fabricate simple thickness mode piezoelectric devices using lithium niobate (LN). Such devices have been shown to atomize liquid ...

Fabrication of Nanoheight Channels Incorporating Surface Acoustic Wave Actuation via Lithium Niobate for Acoustic Nanofluidics

Samenvatting

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