This material is based on research partially sponsored by the United States Office of Naval Research under Grant No. N00014-15-1-2481 and the National Science Foundation under Grant No. 1653396. The views and conclusions contained herein are those of the authors and should not be interpreted as necessarily representing the official policies or endorsements, either expressed or implied, of U.S. Office of Naval Research, the National Science Foundation, or the United States Government.

1. Create the Patterning Map
<ol>
	<li>Using a graphics editor, create the desired pattern for the hemiwicking structures represented as a bitmap image. 
	NOTE: Some of the wicking design parameters (i.e., angle gradient, depth gradient) can be made to be dependent on the grayscale values assigned to each pixel. These grayscale values are then edited in order to modify the desired parameter.</li>
	<li>Save the bitmap as a portable network graphic (.png) and place the file in a readily available folder.</li>
</ol>
2. Placing the Plastic to be Stamped for Molding
<ol>
	<li>Begin by translating the stamping bit away from the workspace to avoid any accidental contact that may cause breakage of the tip (+z displacement, Figure 1).</li>
	<li>Secure the plastic stamping mold/wafer to a backing plate for subsequent stamping on the x,&#160;y translation stage (see Figure 1). Secure the sample/backing plate on to the x, y motorized stamping stage (Figure 1)</li>
	<li>Align the center of the plastic mold/wafer with the stamping axis of the stamping bit. This is accomplished via computerized &#177;x and &#177;y displacements with the x, y motorized stamping stage.</li>
	<li>Translate the stamping bit towards the plastic mold/wafer (-z displacement, Figure 1) until the stamping bit is almost in contact with the mold/wafer surface.</li>
</ol>
3. Stamping the Plastic Sample for PDMS Molding
<ol>
	<li>Using the computerized stamping control program, set the distance between the stamping bit (tip) and the plastic mold/wafer surface.</li>
	<li>Translate the stamping bit in small increments (-&#948;z displacement, Figure 1) towards the surface of the sample until the tooling is in contact with the plastic. 
	NOTE: The bit should only lightly contact the surface.</li>
	<li>After contact, translate the stamping bit away from the sample to avoid any possible contact between the bit and sample during subsequent translation (&#948;z &#8776; 100 &#956;m).</li>
	<li>Assign a pixel distance (in micron), maximum and minimum cavity depth (in micron), maximum and minimum angle (in degrees), initial x and y pixel position of the pattern, and pixel threshold for any gray-scale linked patterning for the stamping procedure.</li>
	<li>Upload the patterning map (created in step 1.1) to be read by the program. Based on the pixel distance and the patterning map, the locations of all the stamps are sent to the stepper motors.</li>
	<li>Ensure that the heating laser is focused on the tip of the stamping bit and only activates while the stamping bit is moving toward and into the plastic mold.</li>
	<li>Create the cavities by pressing the bit into the plastic while following the patterning map to achieve the desired hemiwicking pattern.</li>
	<li>Remove the stamped plastic mold for subsequent surface refinishing and polishing.</li>
	<li>Polish the surface of the plastic mold using 9000 grit, finer wet/dry sandpaper. 
	NOTE: Alternatively, micro-mesh abrasive can be used to ensure the removal of surface deposits that cause cratering around the pillars in the PDMS mold.</li>
</ol>
4. Create the PDMS Molding
<ol>
	<li>Pour 2 g of elastomer base and 0.2 g of the elastomer curing agent into a beaker and mix together thoroughly for 3 min.</li>
	<li>Place the mixture into an evacuated chamber to release any air bubbles caught in the mixture; this step may need to be repeated multiple times. 
	NOTE: For samples of varying volume requirements, adjust the amount of base and curing agent as needed while maintaining a 10:1 ratio.</li>
	<li>Place the stamped plastic mold into a walled container, ideally not much larger than the outer diameter of the mold, for the curing to occur.</li>
	<li>Pour the PDMS mixture free of air pockets onto the stamped plastic and within the container. Pour in a spiral, starting from the center of the stamped area, to attempt to distribute the PDMS mixture as equally as possible.</li>
	<li>Repeat step 4.2 for any air pockets that may have formed from pouring the mixture onto the stamped pattern. Place the PDMS mixture and plastic piece with stamped pattern onto a hot plate and heat the assembly at 100&#160;&#176;C for 15 min. Then heat an additional 25 min at 65&#160;&#176;C.</li>
	<li>Allow the PDMS mixture to cool and cure for 20 min before handling.</li>
	<li>Cut the edges of the PDMS plastic away from the container wall and remove the PDMS plastic from the mold. Store the PDMS plastic in a covered container to avoid dust particles from collecting on the surface.</li>
</ol>
5. Depositing the Thin-Film Metal on the PDMS
<ol>
	<li>Place the sample PDMS inside the deposition chamber leaving enough space for the shutter to be opened and closed unobstructed.</li>
	<li>Depressurize the deposition chamber to at least 10 mTorr.</li>
	<li>Engage the dry pump system and set the spin rate to 75 kRPM. Allow chamber to reach a pressure on the order of 10-8 Torr. 
	NOTE: This will remove most contaminants from the chamber; process may take up to 12 h to complete.</li>
	<li>Power on the cooler and DC power supply and set the power to 55 W.</li>
	<li>Open the argon valve slightly and pressurize the chamber to the order of 10-3 Torr. Set the dry pump system 50 kRPM and wait until this set speed is achieved.</li>
	<li>Reduce power to 35 W and depressurize the chamber to 13 mTorr. Open the shutter to ignited plasma and start the timer. 
	NOTE: Ignited plasma should give off a blue, incandescent glow. Timer should be set for desired thickness of film deposit. It has been determined that for 35 W and pressure of approximately 13 mTorr, a rate of 7 nm deposition per minute is expected.</li>
	<li>Once the desired film thickness has been achieved, close the shutter and turn off power supply.</li>
	<li>Close all of the valves within the deposition chamber and turn off the dry pump system. Allow time for the dry-pump fan to come to a complete stop.</li>
	<li>Slowly pressurize the chamber until it reaches local atmospheric pressure and remove the sample, storing it for future experiments.</li>
</ol>

<ol><li>Plawsky, J. L., et al. <a target="_blank" href="https://www.tandfonline.com/doi/abs/10.1080/15567265.2013.878419">Nano- and Micro-structures for Thin Film Evaporation - A Review.</a> Nanoscale and Microscale Thermophysical Engineering. 18, 251-269 (2014).</li>
<li>Derjaguin, B. V., Churaev, N. V. <a target="_blank" href="https://www.sciencedirect.com/science/article/pii/0021979778900565">On the question of determining the concept of disjoining pressure and its role in the equilibrium and flow of thin films.</a> Journal of Colloid and Interface Science. 66, 389(1978).</li>
<li>Ma, H. B., Cheng, P., Borgmeyer, B., Wang, Y. X. <a target="_blank" href="https://link.springer.com/10.1007%2Fs10404-007-0172-5?LI=true&?from=SL">Fluid flow and heat transfer in the evaporating thin film region.</a> Microfluidics and Nanofluidics. 4 (3), 237-243 (2008).</li>
<li>Hohmann, C., Stephan, P. <a target="_blank" href="https://www.sciencedirect.com/science/article/pii/S089417770200122X">Microscale temperature measurement at an evaporating liquid meniscus.</a> Experimental Thermal and Fluid Science. 26 (2-4), 157-162 (2002).</li>
<li>Potask, M. Jr, Wayner, P. C. Jr <a target="_blank" href="https://www.sciencedirect.com/science/article/pii/0017931072900580">Evaporation from a two-dimensional extended meniscus.</a> International Journal of Heat Mass Transfer. 15 (10), 1851-1863 (1972).</li>
<li>Panchamgam, S. S., Plawsky, J. L., Wayner, P. C. <a target="_blank" href="https://www.sciencedirect.com/science/article/pii/S0894177706000264">Microscale heat transfer in an evaporating moving extended meniscus.</a> Experimental Thermal and Fluid Science. 30 (8), 745-754 (2006).</li>
<li>Arends, A. A., Germain, T. M., Owens, J. F., Putnam, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Simultaneous+Reflectometry+and+Interferometry+for+Measuring+Thin-film+Thickness+and+Curvature%5BTitle%5D)+AND+%22Review+of+Scientific+Instruments%22%5BJournal%5D)">Simultaneous Reflectometry and Interferometry for Measuring Thin-film Thickness and Curvature.</a> Review of Scientific Instruments. 89 (5), (2018).</li>
<li>Zhu, Y., Antao, D. S., Lu, Z., Somasundaram, S., Zhang, T., Wang, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Prediction+and+characterization+of+dry+out+heat+flux+in+micropillar+wick+structures%5BTitle%5D)+AND+%22Langmuir%22%5BJournal%5D)">Prediction and characterization of dry out heat flux in micropillar wick structures.</a> Langmuir. 32 (7), 1920-1927 (2016).</li>
<li>Kim, J., Moon, M. W., Kim, H. Y. <a target="_blank" href="https://www.cambridge.org/core/journals/journal-of-fluid-mechanics/article/dynamics-of-hemiwicking/E42C2F820D7D4EE9BF9CCEA0A7162AAA">Dynamics of hemiwicking.</a> Journal of Fluid Mechanics. 800, 57-71 (2016).</li>
<li>Ding, C., Soni, G., Bozorgi, P., Meinhart, C. D., MacDonald, N. C. <a target="_blank" href="https://www.nsti.org/publications/Nanotech/2009/pdf/706.pdf">Wicking Study of Nanostructured Titania Surfaces for Flat Heat Pipes.</a> Nanotech Conference & Expo. , Houston, TX. (2009).</li>
<li>Chen, R., Lu, M. C., Srinivasan, V., Wang, Z., Cho, H. H., Majumdar, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Nanowires+for+Enhanced+Boiling+Heat+Transfer%5BTitle%5D)+AND+%22Nano+Letters%22%5BJournal%5D)">Nanowires for Enhanced Boiling Heat Transfer.</a> Nano Letters. 9 (2), 548-553 (2009).</li>
<li>Kim, B. S., Choi, G., Shim, D. II, Kim, K. M., Cho, H. H. <a target="_blank" href="https://www.sciencedirect.com/science/article/pii/S0017931015306591">Surface roughening for hemi-wicking and its impact on convective boiling heat transfer.</a> International Journal of Heat and Mass Transfer. 102, 1100-1107 (2016).</li>
<li>Mikkelsen, M. B., et al. <a target="_blank" href="http://iopscience.iop.org/article/10.1088/0960-1317/21/11/115008/meta">Controlled deposition of sol-gel sensor material using hemiwicking.</a> Journal of Micromechanics and Microengineering. 21 (11), (2011).</li>
<li>Haatainen, T., Ahopelto, J. <a target="_blank" href="http://iopscience.iop.org/article/10.1238/Physica.Regular.067a00357/meta">Pattern Transfer using Step&Stamp Imprint Lithography.</a> Physica Scripta. 67 (4), 357-360 (2003).</li>
<li>Chou, S. Y., Krauss, P. R., Renstrom, P. J. <a target="_blank" href="http://www.nanonex.com/nanoimprint.pdf">Nanoimprint lithography.</a> Journal of vacuum Science & Technology B: Microelectronics and Nanometer Structures Processing, Measurement, and Phenomena. 14 (6), 4129(1996).</li>
<li>Pozzato, A., et al. <a target="_blank" href="https://www.sciencedirect.com/science/article/abs/pii/S0167931706000268">Superhydrophobic surfaces fabricated by nanoprint lithography.</a> Microelectronic Engineering. 83 (4-9), 884-888 (2006).</li>
<li>Nair, R. P., Zou, M. <a target="_blank" href="https://www.sciencedirect.com/science/article/pii/S025789720800652X">Surface-nano-texturing by aluminum-induced crystallization of amorphous silicon.</a> Surface and Coatings Technology. 203 (5-7), 675-679 (2008).</li>
<li>Ashby, P. D., Lieber, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Ultra-sensitive+Imaging+and+Interfacial+Analysis+of+Patterned+Hydrophilic+SAM+Surfaces+Using+Energy+Dissipation+Chemical+Force+Microscopy%5BTitle%5D)+AND+%22Journal+of+the+American+Chemical+Society%22%5BJournal%5D)">Ultra-sensitive Imaging and Interfacial Analysis of Patterned Hydrophilic SAM Surfaces Using Energy Dissipation Chemical Force Microscopy.</a> Journal of the American Chemical Society. 127 (18), 6814-6818 (2005).</li>
</ol>

The authors have no disclosures to mention for this paper.

A method has been introduced to create patterned pillar arrays for hemiwicking structures; this is accomplished by imprinting cavities on a plastic wafer with an engraving apparatus that follows patterning from a bitmap created by the user. A PDMS mixture is then poured, cured and coated with a thin film of aluminum via deposition. The pillar array characteristics can be customized depending on the gray scale value that is assigned in the bitmap following this protocol. This crucial aspect of patterning can crea...

Recently there has been increased interest in being able to both actively and passively control the wetting, evaporation, and mixing of fluids. Uniquely textured hemiwicking surfaces provide a novel solution for cooling techniques because these textured surfaces act as a fluid (and/or heat) pump without the moving parts. This fluid motion is driven by a cascade of capillary action events associated with the dynamic curvature of the liquid thin-film. In general, when a fluid wets a solid surface, a curved liquid thin-film (i.e., liquid meniscus) rapidly forms. The fluid thickness and curvature profile evolve until a free-energy minimum is reached. For reference, this dynamic wetting profile can rapidly decay to tens of nanometers in thickness within a spanning (fluid-wetting) length-scale of only tens of micrometers. Thus, this transitional (liquid-film) region can undergo significant changes in liquid-interface curvature. The transitional (thin-film) region is where nearly all the dynamic physics and chemistry originates. In particular, the transitional (thin-film) region is where maximum (1) evaporation rates, (2) dis-joining pressure gradients, and (3) hydrostatic pressure gradients are found1,2. As a result, curved liquid-films play a vital role in thermal transport, phase separation, fluid instabilities, and the mixing of multi-component fluids. For instance, with respect to heat transfer, the highest wall heat fluxes have been observed in this highly curved, transitional thin-film region3,4,5,6,7.
Recent hemiwicking studies have shown that the geometry (e.g., height, diameter, etc.) and placement of the pillars determine the wetting front profile and velocity of the fluid running through the structures8. As the fluid front is evaporating off the end of the last structure in an array, the fluid front is maintained at a constant distance and curvature, as the evaporated fluid is being replaced by the fluid stored in the wicking structures9. Hemiwicking structures have also been used in heat pipes and on boiling surfaces to analyze and enhance different heat transfer mechanisms.10,11,12.
One method currently used to create wicking structures is thermal imprint lithography13. This method is performed by stamping the desired layout into a resist layer on a silicon mold sample with a thermoplastic polymer stamp, then removing the stamp to maintain the microstructures. Once removed, the sample is put through a reactive ion etching process to remove any of the excess resist layer14,15. This process, however, can be sensitive to the temperature of the fabrication of the wicking structures and includes multiple steps that utilize various coatings to ensure the accuracy of the wicking structures16. It is also the case that lithography techniques are not practical for macro-scale patterning; while they still provide a way to create a pattern of microstructures on a surface, the throughput of this procedure is far less than ideal for large-scale reproduction. Considering large-scale, reproducible texturing, such as spin or dip coating, there is an inherent lack of controllable patterning. These methods create a random array of microstructures on the target surface but can be scaled to cover vastly larger areas than traditional lithography techniques17.
The protocol outlined within this report attempts to combine the strengths of traditional texturing methods while simultaneously eliminating the specific weaknesses of each; it defines a way to fabricate custom hemiwicking structures of various heights, shapes, orientations, and materials on a macro-scale and with potentially high throughput. Various wicking patterns can be quickly created for the purpose of optimization of wicking characteristics, such as directional control of fluid velocity, propagation, and mixing of different fluids. The use of different wicking structures can also provide varying thin-film thickness and curvature profiles, which can be used to systematically study the coupling between heat and mass transfer with different thickness and curvature profiles of the liquid meniscus.

<table><tbody><tr><td>NI-DAQ 9403</td><td>National Instruments</td><td>370466AE-01</td><td>The communication interface between the camera and the control switch for the laser.</td></tr><tr><td>Control Switch</td><td>Crouzet</td><td>GN84134750</td><td>A controller to use for the laser that activates the laser based on the voltage sent by the DAQ.</td></tr><tr><td>Flea Camera</td><td>FLIR</td><td>FL3-U3-120S3C-C</td><td>A flea camera used for imaging the drill bit on the plastic mold.&amp;nbsp;</td></tr><tr><td>Flea Imaging Camera</td><td>Point Grey</td><td>FL3-U3-20E4M-C</td><td>A flea camera used for obtaining the side images of the pillars.</td></tr><tr><td>200 Steps/rev, 12V-350mA Stepper Motor (x2)</td><td>AdaFruit</td><td>324</td><td>The stepper motors are used to control the depth and angle of the end mill.&amp;nbsp;</td></tr><tr><td>10x Infinity Corrected Long Working Distance Objective</td><td>Mitutoyo&amp;nbsp;</td><td>#46-144</td><td>The objective used to get the image of the side of the pillars.</td></tr><tr><td>15x Infinite Conjugate, UV Coated, ReflX Objective</td><td>TechSpec</td><td>#58-417</td><td>The objective used to get the image of the top of the pillars.&amp;nbsp;</td></tr><tr><td>72002 0.002D X 0.006 LOC Carbide SQ 2FL Miniature End Mill</td><td>Harvey Tools</td><td>72002</td><td>The drill bit that was used to create holes in the plastic mold.&amp;nbsp;</td></tr><tr><td>DC Power Delivery at 1 kW</td><td>Advanced Energy</td><td>MDX-1K</td><td>Used to power the deposition sputterer.&amp;nbsp;</td></tr><tr><td>Turbo-V 70LP Nacro Torr Pump</td><td>Varian</td><td>9699336</td><td>Turbo Pump used to reduce pressure inside deposition chamber.</td></tr><tr><td>2000mw, 405nm High-Power Blue Light Focus Laser</td><td>WDLasers</td><td>KREE</td><td>Sample Heating Laser</td></tr><tr><td>5.875&quot; I.D. Dessicator w/ 0.25&quot; Tube Connections</td><td>McMaster-Carr</td><td>2204K5</td><td>PDMS Dessicator</td></tr><tr><td>SYLGARD 184 Silicone Elastomer, 0.5kg Kit</td><td>Dow-Corning</td><td>4019862</td><td>The PDMS Kit used to make the base.</td></tr><tr><td>Diaphragm Air Compressor / Vacuum Pump</td><td>Gast</td><td>DOL-701-AA</td><td>Dessicator Vacuum Pump</td></tr><tr><td>Motorized Linear Stages (2x)</td><td>Standa</td><td>8MT175</td><td>The stepper motors used to control the sample plate in the x- and y- direction.&amp;nbsp;</td></tr><tr><td>2&quot; Diameter Unmounted Poistive Achromatic Doublets, AR Coated: 400-700 nm</td><td>ThorLabs</td><td>AC508-150-A</td><td>The achromat was ued in order to obtain the images of the side of the pillars.&amp;nbsp;</td></tr><tr><td>Flea 3 Mono&amp;nbsp; Camera, 2448 X 2048 Pixels</td><td>Point Grey</td><td>FL3-GE-50S5M-C</td><td>A flea camera used for imiaging the top of the pillars.</td></tr><tr><td>Digital Vacuum Transducer</td><td>Thyrcont Vacuum Instruments</td><td>4940-CF-212734</td><td>Used for monitoring pressure inside deposition chamber.</td></tr><tr><td>Pressurized Argon Tank Resovoir</td><td>Airgas</td><td>AR RP300</td><td>Gas used in deposition process.</td></tr><tr><td>1-D Translation Stage</td><td>Newport Corporation</td><td>TSX-1D</td><td>A translation stage used to move the camera to focus on the end mill.&amp;nbsp;</td></tr><tr><td>Cylindrical Laser Mount (x2)</td><td>Newport Corporation</td><td>ULM-TILT-M</td><td>The laser mount was used to move the camera to focus on the end mill.</td></tr><tr><td>Benchtop Chiller with Centrifugal Pump, 120V, 60Hz</td><td>Polyscience</td><td>LS51MX1A110C</td><td>A chiller used for the deposition assembly.</td></tr><tr><td>Alcatel Adixen 2010SD XP, Explosion Proof Motor, Rotary Vane Vacuum Pump, 1-Phase</td><td>Ideal Vacuum Products</td><td>210SDMLAM-XP</td><td>A vacuum pump used for the deposition assembly.&amp;nbsp;</td></tr><tr><td>Fan, 105 CFM, 115 V (x2)</td><td>Comair Rotron</td><td>MU2A1</td><td>A fan used for cooling certain aspects of the deposition assembly.</td></tr></tbody></table>

scalable stamp printing and fabrication of hemiwicking surfaces

Hemiwicking is a process where a fluid wets a patterned surface beyond its normal wetting length due to a combination of capillary action and imbibition. This wetting phenomenon is important in many technical fields ranging from physiology to aerospace engineering. Currently, several different techniques exist for fabricating hemiwicking structures. These conventional methods, however, are often time consuming and are difficult to scale-up for large areas or are difficult to customize for specific, nonhomogeneous patterning geometries. The presented protocol provides researchers with a simple, scalable, and cost-effective method for fabricating micro-patterned hemiwicking surfaces. The method fabricates wicking structures through the use of stamp printing, polydimethylsiloxane (PDMS) molding, and thin-film surface coatings. The protocol is demonstrated for hemiwicking with ethanol on PDMS micropillar arrays coated with a 70 nm thick aluminum thin-film.

Figure 1&#160;provides a schematic of how the stamping mechanism would create the mold for the wicking structures on a plastic mold. To investigate the quality of the stamping apparatus in manufacturing wicking films, two different pillar arrays were created to analyze the quality of the pillars for future wicking experiments. Aspects of the apparatus investigated were the accuracy of the height of the pillars (with and without a depth gradient), the quality ...

Watch this Scientific Journal Video about Scalable Stamp Printing and Fabrication of Hemiwicking Surfaces at JoVE.com

Scalable Stamp Printing and Fabrication of Hemiwicking Surfaces

A simple protocol is provided for the fabrication of hemiwicking structures of varying sizes, shapes, and materials. The protocol uses a combination of physical stamping, PDMS molding, and thin-film surface modifications via common materials deposition techniques.

Hemiwicking is a process where a fluid wets a patterned surface beyond its normal wetting length due to a combination of capillary action and imbibition. ...

scalable-stamp-printing-and-fabrication-of-hemiwicking-surfaces

Research

JoVE Journal

Engineering

7.3K Views.  University of Central Florida. A simple protocol is provided for the fabrication of hemiwicking structures of varying sizes, shapes, and materials. The protocol uses a combination of physical stamping, PDMS molding, and thin-film surface modifications via common materials deposition techniques. 

Video: Scalable Stamp Printing and Fabrication of Hemiwicking Surfaces

Micropatterned Surfaces to Study Hyaluronic Acid Interactions with Cancer Cells

Cancer invasion and progression involves a motile cell phenotype, which is under complex regulation by growth factors/cytokines and extracellular matrix (ECM) components within the tumor microenvironment. Hyaluronic acid (HA) is one stromal ECM component that is known to facilitate tumor progression by enhancing invasion, growth, and angiogenesis1. Interaction of HA with its cell surface receptor CD44 induces signaling events that promote tumor cell growth, survival, and migration, thereby increasing metastatic spread2-3. HA is an anionic, nonsulfated glycosaminoglycan composed of repeating units of D-glucuronic acid and D-N-acetylglucosamine. Due to the presence of carboxyl and hydroxyl groups on repeating disaccharide units, native HA is largely hydrophilic and amenable to chemical modifications that introduce sulfate groups for photoreative immobilization 4-5. Previous studies involving the immobilizations of HA onto surfaces utilize the bioresistant behavior of HA and its sulfated derivative to control cell adhesion onto surfaces6-7. In these studies cell adhesion preferentially occurs on non-HA patterned regions.
To analyze cellular interactions with exogenous HA, we have developed patterned functionalized surfaces that enable a controllable study and high-resolution visualization of cancer cell interactions with HA. We utilized microcontact printing (uCP) to define discrete patterned regions of HA on glass surfaces. A "tethering" approach that applies carbodiimide linking chemistry to immobilize HA was used 8. Glass surfaces were microcontact printed with an aminosilane and reacted with a HA solution of optimized ratios of EDC and NHS to enable HA immobilization in patterned arrays. Incorporating carbodiimide chemistry with mCP enabled the immobilization of HA to defined regions, creating surfaces suitable for in vitro applications. Both colon cancer cells and breast cancer cells implicitly interacted with the HA micropatterned surfaces. Cancer cell adhesion occurred within 24 hours with proliferation by 48 hours. Using HA micropatterned surfaces, we demonstrated that cancer cell adhesion occurs through the HA receptor CD44. Furthermore, HA patterned surfaces were compatible with scanning electron microscopy (SEM) and allowed high resolution imaging of cancer cell adhesive protrusions and spreading on HA patterns to analyze cancer cell motility on exogenous HA.

A novel approach that allows the high-resolution analysis of cancer cell interactions with exogenous hyaluronic acid (HA) is described. Patterned surfaces are fabricated by combining carbodiimide chemistry and microcontact printing.

Cancer invasion and progression involves a motile cell phenotype, which is under complex regulation by growth factors/cytokines and extracellular matrix ...

Bioengineering

Creating Two-Dimensional Patterned Substrates for Protein and Cell Confinement

Microcontact printing provides a rapid, highly reproducible method for the creation of well-defined patterned substrates.1 While microcontact printing can be employed to directly print a large number of molecules, including proteins,2 DNA,3 and silanes,4 the formation of self-assembled monolayers (SAMs) from long chain alkane thiols on gold provides a simple way to confine proteins and cells to specific patterns containing adhesive and resistant regions. This confinement can be used to control cell morphology and is useful for examining a variety of questions in protein and cell biology. Here, we describe a general method for the creation of well-defined protein patterns for cellular studies.5 This process involves three steps: the production of a patterned master using photolithography, the creation of a PDMS stamp, and microcontact printing of a gold-coated substrate. Once patterned, these cell culture substrates are capable of confining proteins and/or cells (primary cells or cell lines) to the pattern.
The use of self-assembled monolayer chemistry allows for precise control over the patterned protein/cell adhesive regions and non-adhesive regions; this cannot be achieved using direct protein stamping. Hexadecanethiol, the long chain alkane thiol used in the microcontact printing step, produces a hydrophobic surface that readily adsorbs protein from solution. The glycol-terminated thiol, used for backfilling the non-printed regions of the substrate, creates a monolayer that is resistant to protein adsorption and therefore cell growth.6 These thiol monomers produce highly structured monolayers that precisely define regions of the substrate that can support protein adsorption and cell growth. As a result, these substrates are useful for a wide variety of applications from the study of intercellular behavior7 to the creation of microelectronics.8
While other types of monolayer chemistry have been used for cell culture studies, including work from our group using trichlorosilanes to create patterns directly on glass substrates,9 patterned monolayers formed from alkane thiols on gold are straight-forward to prepare. Moreover, the monomers used for monolayer preparation are commercially available, stable, and do not require storage or handling under inert atmosphere. Patterned substrates prepared from alkane thiols can also be recycled and reused several times, maintaining cell confinement.10

Self-assembled monolayers (SAMs) formed from long chain alkane thiols on gold provide well-defined substrates for the formation of protein patterns and cell confinement. Microcontact printing of hexadecanethiol using a polydimethylsiloxane (PDMS) stamp followed by backfilling with a glycol-terminated alkane thiol monomer produces a pattern where protein and cells adsorb only to the stamped hexadecanethiol region.

Microcontact printing provides a rapid, highly reproducible method for the creation of well-defined patterned substrates.1  While microcontact printing ...

Micro-masonry for 3D Additive Micromanufacturing

Transfer printing is a method to transfer solid micro/nanoscale materials (herein called &#8216;inks&#8217;) from a substrate where they are generated to a different substrate by utilizing elastomeric stamps. Transfer printing enables the integration of heterogeneous materials to fabricate unexampled structures or functional systems that are found in recent advanced devices such as flexible and stretchable solar cells and LED arrays. While transfer printing exhibits unique features in material assembly capability, the use of adhesive layers or the surface modification such as deposition of self-assembled monolayer (SAM) on substrates for enhancing printing processes hinders its wide adaptation in microassembly of microelectromechanical system (MEMS) structures and devices. To overcome this shortcoming, we developed an advanced mode of transfer printing which deterministically assembles individual microscale objects solely through controlling surface contact area without any surface alteration. The absence of an adhesive layer or other modification and the subsequent material bonding processes ensure not only mechanical bonding, but also thermal and electrical connection between assembled materials, which further opens various applications in adaptation in building unusual MEMS devices.

This paper introduces a 3D additive micromanufacturing strategy (termed &#8216;micro-masonry&#8217;) for the flexible fabrication of microelectromechanical system (MEMS) structures and devices. This approach involves transfer printing-based assembly of micro/nanoscale materials in conjunction with rapid thermal annealing-enabled material bonding techniques.

Transfer printing is a method to transfer solid micro/nanoscale materials (herein called &#8216;inks&#8217;) from a substrate where they are generated to ...

Integration of Light Trapping Silver Nanostructures in Hydrogenated Microcrystalline Silicon Solar Cells by Transfer Printing

One of the potential applications of metal nanostructures is light trapping in solar cells, where unique optical properties of nanosized metals, commonly known as plasmonic effects, play an important role. Research in this field has, however, been impeded owing to the difficulty of fabricating devices containing the desired functional metal nanostructures. In order to provide a viable strategy to this issue, we herein show a transfer printing-based approach that allows the quick and low-cost integration of designed metal nanostructures with a variety of device architectures, including solar cells. Nanopillar poly(dimethylsiloxane) (PDMS) stamps were fabricated from a commercially available nanohole plastic film as a master mold. On this nanopatterned PDMS stamps, Ag films were deposited, which were then transfer-printed onto block copolymer (binding layer)-coated hydrogenated microcrystalline Si (&#181;c-Si:H) surface to afford ordered Ag nanodisk structures. It was confirmed that the resulting Ag nanodisk-incorporated &#181;c-Si:H solar cells show higher performances compared to a cell without the transfer-printed Ag nanodisks, thanks to plasmonic light trapping effect derived from the Ag nanodisks. Because of the simplicity and versatility, further device application would also be feasible thorough this approach.

A viable transfer printing-based methodology to introduce plasmonic metal nanostructures in solar cells is described. Using nanopillar poly(dimethylsiloxane) stamps, an Ag-based ordered nanodisk array was integrated with standard hydrogenated microcrystalline Si solar cells, which led to improved device performances due to plasmonic light trapping.

One of the potential applications of metal nanostructures is light trapping in solar cells, where unique optical properties of nanosized metals, commonly ...

Creating Adhesive and Soluble Gradients for Imaging Cell Migration with Fluorescence Microscopy

Cells can sense and migrate towards higher concentrations of adhesive cues such as the glycoproteins of the extracellular matrix and soluble cues such as growth factors. Here, we outline a method to create opposing gradients of adhesive and soluble cues in a microfluidic chamber, which is compatible with live cell imaging. A copolymer of poly-L-lysine and polyethylene glycol (PLL-PEG) is employed to passivate glass coverslips and prevent non-specific adsorption of biomolecules and cells. Next, microcontact printing or dip pen lithography are used to create tracks of streptavidin on the passivated surfaces to serve as anchoring points for the biotinylated peptide arginine-glycine-aspartic acid (RGD) as the adhesive cue. A microfluidic device is placed onto the modified surface and used to create the gradient of adhesive cues (100% RGD to 0% RGD) on the streptavidin tracks. Finally, the same microfluidic device is used to create a gradient of a chemoattractant such as fetal bovine serum (FBS), as the soluble cue in the opposite direction of the gradient of adhesive cues.

A method for the assembly of adhesive and soluble gradients in a microscopy chamber for live cell migration studies is described. The engineered environment combines antifouling surfaces and adhesive tracks with solution gradients and therefore allows one to determine the relative importance of guidance cues.

Cells can sense and migrate towards higher concentrations of adhesive cues such as the glycoproteins of the extracellular matrix and soluble cues such as ...

Fabricating Complex Culture Substrates Using Robotic Microcontact Printing (R-&#181;CP) and Sequential Nucleophilic Substitution

In tissue engineering, it is desirable to exhibit spatial control of tissue morphology and cell fate in culture on the micron scale. Culture substrates presenting grafted poly(ethylene glycol) (PEG) brushes can be used to achieve this task by creating microscale, non-fouling and cell adhesion resistant regions as well as regions where cells participate in biospecific interactions with covalently tethered ligands. To engineer complex tissues using such substrates, it will be necessary to sequentially pattern multiple PEG brushes functionalized to confer differential bioactivities and aligned in microscale orientations that mimic in vivo niches. Microcontact printing (&#956;CP) is a versatile technique to pattern such grafted PEG brushes, but manual &#956;CP cannot be performed with microscale precision. Thus, we combined advanced robotics with soft-lithography techniques and emerging surface chemistry reactions to develop a robotic microcontact printing (R-&#956;CP)-assisted method for fabricating culture substrates with complex, microscale, and highly ordered patterns of PEG brushes presenting orthogonal &#8216;click&#8217; chemistries. Here, we describe in detail the workflow to manufacture such substrates.

Fabricating Complex Culture Substrates Using Robotic Microcontact Printing R- &#181;CP and Sequential Nucleophilic Substitution

Cell culture substrates functionalized with microscale patterns of biological ligands have immense utility in the field of tissue engineering. Here, we demonstrate the versatile and automated manufacture of tissue culture substrates with multiple, micropatterned poly(ethylene glycol) brushes presenting orthogonal chemistries that enable spatially precise and site-specific immobilization of biological ligands.

In tissue engineering, it is desirable to exhibit spatial control of tissue morphology and cell fate in culture on the micron scale. Culture substrates ...

Fabricating Reactive Surfaces with Brush-like and Crosslinked Films of Azlactone-Functionalized Block Co-Polymers

In this paper, fabrication methods that generate novel surfaces using the azlactone-based block co-polymer, poly (glycidyl methacrylate)-block-poly (vinyl dimethyl azlactone) (PGMA-b-PVDMA), are presented. Due to the high reactivity of azlactone groups towards amine, thiol, and hydroxyl groups, PGMA-b-PVDMA surfaces can be modified with secondary molecules to create chemically or biologically functionalized interfaces for a variety of applications. Previous reports of patterned PGMA-b-PVDMA interfaces have used traditional top-down patterning techniques that generate non-uniform films and poorly controlled background chemistries. Here, we describe customized patterning techniques that enable precise deposition of highly uniform PGMA-b-PVDMA films in backgrounds that are chemically inert or that have biomolecule-repellent properties. Importantly, these methods are designed to deposit PGMA-b-PVDMA films in a manner that completely preserves azlactone functionality through each processing step. Patterned films show well-controlled thicknesses that correspond to polymer brushes (~90 nm) or to highly crosslinked structures (~1-10 &#956;m). Brush patterns are generated using either the parylene lift-off or interface directed assembly methods described and are useful for precise modulation of overall chemical surface reactivity by adjusting either the PGMA-b-PVDMA pattern density or the length of the VDMA block. In contrast, the thick, crosslinked PGMA-b-PVDMA patterns are obtained using a customized micro-contact printing technique and offer the benefit of higher loading or capture of secondary material due to higher surface area to volume ratios. Detailed experimental steps, critical film characterizations, and trouble-shooting guides for each fabrication method are discussed.

Surface fabrication methods for patterned deposition of nanometer thick brushes or micron thick, crosslinked films of an azlactone block co-polymer are reported. Critical experimental steps, representative results, and limitations of each method are discussed. These methods are useful for creating functional interfaces with tailored physical features and tunable surface reactivity.

In this paper, fabrication methods that generate novel surfaces using the azlactone-based block co-polymer, poly (glycidyl methacrylate)-block-poly (vinyl ...

Hybrid Printing for the Fabrication of Smart Sensors

A method to combine additively manufactured substrates or foils and multilayer inkjet printing for the fabrication of sensor devices is presented. First, three substrates (acrylate, ceramics, and copper) are prepared. To determine the resulting material properties of these substrates, profilometer, contact angle, scanning electron microscope (SEM), and focused ion beam (FIB) measurements are done. The achievable printing resolution and suitable drop volume for each substrate are, then, found through the drop size tests. Then, layers of insulating and conductive ink are inkjet printed alternately to fabricate the target sensor structures. After each printing step, the respective layers are individually treated by photonic curing. The parameters used for the curing of each layer are adapted depending on the printed ink, as well as on the surface properties of the respective substrate. To confirm the resulting conductivity and to determine the quality of the printed surface, four-point probe and profilometer measurements are done. Finally, a measurement set-up and results achieved by such an all-printed sensor system are shown to demonstrate the achievable quality.

Here we present a protocol for the fabrication of inkjet-printed multilayer sensor structures on additively manufactured substrates and foil.

A method to combine additively manufactured substrates or foils and multilayer inkjet printing for the fabrication of sensor devices is presented. First, ...

Automated Robotic Dispensing Technique for Surface Guidance and Bioprinting of Cells

This manuscript describes the introduction of cell guidance features followed by the direct delivery of cells to these features in a hydrogel bioink using an automated robotic dispensing system. The particular bioink was selected as it allows cells to sediment towards and sense the features. The dispensing system bioprints viable cells in hydrogel bioinks using a backpressure assisted print head. However, by replacing the print head with a sharpened stylus or scalpel, the dispensing system can also be employed to create topographical cues through surface etching. The stylus movement can be programmed in steps of 10 &#181;m in the X, Y and Z directions. The patterned grooves were able to orientate mesenchymal stem cells, influencing them to adopt an elongated morphology in alignment with the grooves&#39; direction. The patterning could be designed using plotting software in straight lines, concentric circles, and sinusoidal waves. In a subsequent procedure, fibroblasts and mesenchymal stem cells were suspended in a 2% gelatin bioink, for bioprinting in a backpressure driven extrusion printhead. The cell bearing bioink was then printed using the same programmed coordinates used for the etching. The bioprinted cells were able to sense and react to the etched features as demonstrated by their elongated orientation along the direction of the etched grooves.

This protocol describes a bioprinting methodology using an automated robotic depositing system that incorporates etched topographical guidance cues with the precision deposition of a cell bearing hydrogel bioink. The printed cells are directly delivered to the etched features and are able to sense and orientate with them.

This manuscript describes the introduction of cell guidance features followed by the direct delivery of cells to these features in a hydrogel bioink using ...

Preparation of Tunable Extracellular Matrix Microenvironments to Evaluate Schwann Cell Phenotype Specification

Traumatic peripheral nervous system (PNS) injuries currently lack suitable treatments to regain full functional recovery. Schwann cells (SCs), as the major glial cells of the PNS, play a vital role in promoting PNS regeneration by dedifferentiating into a regenerative cell phenotype following injury. However, the dedifferentiated state of SCs is challenging to maintain through the time-period needed for regeneration and is impacted by changes in the surrounding extracellular matrix (ECM). Therefore, determining the complex interplay between SCs and differing ECM to provide cues of regenerative potential of SCs is essential. To address this, a strategy was created where different ECM proteins were adsorbed onto a tunable polydimethylsiloxane (PDMS) substrate which provided a platform where stiffness and protein composition can be modulated. SCs were seeded onto the tunable substrates and critical cellular functions representing the dynamics of SC phenotype were measured. To illustrate the interplay between SC protein expression and cellular morphology, differing seeding densities of SCs in addition to individual microcontact printed cellular patterns were utilized and characterized by immunofluorescence staining and western blot. Results showed that cells with a smaller spreading area and higher extent of cellular elongation promoted higher levels of SC regenerative phenotypic markers. This methodology not only begins to unravel the significant relationship between the ECM and cellular function of SCs, but also provides guidelines for the future optimization of biomaterials in peripheral nerve repair.

This methodology aims to illustrate the mechanisms by which extracellular matrix cues such as substrate stiffness, protein composition and cell morphology regulate Schwann cell (SC) phenotype.

Traumatic peripheral nervous system (PNS) injuries currently lack suitable treatments to regain full functional recovery. Schwann cells (SCs), as the ...