1. Preparing the microfluidic flow cell
NOTE: For this protocol, a commercial microfluidic flow cell will be utilized. By using a commercial product that is designed for light penetration from an optical microscope, any challenges regarding brightfield illumination of the media will be minimized.
<ol>
	<li>Start preparing the microfluidic flow cell by covering the outlet with parafilm to seal one end of the channel so that the empty flow cell may be packed with polymeric powder. Before starting the experiment, confirm that the microfluidic channel is clean and dry.
	<ol>
		<li>Tape the metric paper ruler directly beneath the flow channel.</li>
		<li>Weigh the microfluidic flow cell with the parafilm and the ruler attached. The mass of the flow cell is the unpacked flow cell mass (mu).</li>
	</ol>
	</li>
</ol>
2. Packing the powder into the channel
<ol>
	<li>When packing the powder, use a plastic pipet to transfer the powder. Note that particles may adhere to the outside of the pipet tip, which is a result of tribocharging.
	<ol>
		<li>While introducing the powder into the channel, tap the flow cell at least five times to compact the powder. Continue packing until the powder reaches the beginning of the opening of the flow channel. 
		NOTE: Tapping compacts the powder within the channel with the goal of providing a reproducible diagnostic tool. For certain applications, this effort can be a higher, lower, or equivalent level of powder compaction than the compaction that is observed in the application of interest. If there are issues with the reproducibility of tapping or with packing powder within the application, consider performing ASTM D7481-1820.</li>
		<li>Remove the powder present on the outer surface of the flow cell with a wipe soaked in alcohol. 
		NOTE: Some types of particles can be hydrophobic, so water may not remove the particles well.</li>
	</ol>
	</li>
	<li>Once the powders are packed, visually inspect the flow cell for loosely packed powder. If the powder within the flow cell appears loosely packed (Figure 1), tap the flow cell five more times. If the powder packing appears consistent and compact, weigh the flow cell to measure the mass of the polymeric powder (mp - mu; see Equation 1).
	<ol>
		<li>Calculate the bulk packing density (&#961;) using the difference between the unpacked (mu) and packed flow cell mass (mp) and dividing it by the volume of the flow cell. The volume of the flow cell is then known [length (l): 50 mm, width (w): 5 mm, channel depth (h): 0.8 mm]. 
		<img alt="Equation 1" src="/files/ftp_upload/63494/63494eq01.jpg" style="margin: auto;" />&#160; &#160; &#160;Eq 1</li>
		<li>Confirm that the packing density is in the typical range of 0.45 g/mL to 0.55 g/mL for polymeric powders2,3,4,21. Leave the flow cells in the fume hood until steps 3 and 4 are completed. 
		CAUTION: Particles with a diameter of less than 10 &#181;m can penetrate into the lungs and potentially enter the bloodstream, which can cause health problems related to the pulmonary and cardiovascular systems. The polymeric powders that were used in this experiment have a particle diameter of approximately 50 &#956;m. Therefore, inhalation of the particles has less potential to cause health problems, but smaller particles are present even in narrow particle size distributions. For the safest environment, preparation of the flow cells should be done in a fume hood.</li>
	</ol>
	</li>
</ol>
3. Preparing the solvent
<ol>
	<li>Prepare a 75 wt% solution of ethanol in water. Note that the solvent will be referred to as the fluid in the rest of this manuscript. 
	&#8203;CAUTION: Make sure that the beaker used to prepare the solution is free from any surfactants, as surfactants will affect the results.</li>
</ol>
4. Preparing the white light table
<ol>
	<li>To prevent flooding the detector (camera) with too much light, cover the light table with an opaque material, such as a 3D printed cover in black polylactic acid (PLA) filament (Supplementary Figure 1). Ensure that the material has an opening that is the size of the microchannel (5 mm x 55 mm) to allow light to illuminate the powder. 
	NOTE: Too much light means the screen or monitor of the camera will appear white and the microchannel will not be visible. Therefore, the detector will be unable to focus the lens on the microchannel.</li>
	<li>To ensure that the camera on the mobile device can capture the contrast between the wet and dry powder, use the light table at a low to medium light intensity. 
	NOTE: High light intensity is at 100%. The other two settings are relative to the high light intensity; the setting for low light intensity is at ~30%, and medium light intensity is at ~65%.</li>
	<li>Align the camera on the mobile device directly above the light table. Confirm that the camera is perpendicular to the top of the light table (Figure 2).</li>
	<li>Orient the camera on the mobile device so that the long axis of the mobile device aligns with the longest axis of the flow cell.</li>
</ol>
5. Starting the experiment
<ol>
	<li>Place the flow cell on the light table and focus the camera on the mobile device on the flow channel. 
	NOTE: For optimal results, a darker (reduced overhead lighting) recording space will typically provide better image resolution. If a dark space is not available, minimizing changes in overhead lighting (lights being turned on, turned off, or being dimmed) during recording should improve graphic signals and minimize undesired noise in the experiment.</li>
	<li>After focusing the camera on the mobile device, select the record button. Add 125 &#956;L of fluid to the open inlet of the microchannel using a pipet.</li>
	<li>Record the flow for 2 min or until all the powder is wetted visibly.</li>
</ol>
6. Analyzing the data
<ol>
	<li>Transfer the video file from the mobile device to the computer for easy access. Note that videos over 2 min may not load in the software at this time, as the file size can be excessively large.</li>
	<li>Download Tracker, a free software from the Physlets website22. This software can track position, velocity, and acceleration in the following video files: .mov, .avi, .mp4, .flv, .wmv, etc. For the following steps, please refer to Supplemental File. 
	NOTE: For Mac users, install the latest version of the software for the software to function properly. Additionally, Mac users may require a video engine (Xuggle), animated GIF files (.gif), or image sequences that consist of one or more digital images (.jpg, .png, or pasted from the clipboard).</li>
	<li>Once the software is installed, open the Tracker software. From the File menu, select Open File to load the transferred video file from step 6.1 on the desktop of the computer.</li>
	<li>Click the Clip Settings icon, which looks like the film strip, to define the Starting Frame and the Step Size. 
	NOTE: Placing the mouse over an icon will identify the icon.
	<ol>
		<li>Define the Starting Frame. The Starting Frame is defined as the frame in which the first contrast (the contrast between the wet and dry powder) is observed.</li>
		<li>Set the Step Size. Step Size refers to the frame step size, which the software would analyze. From prior experiments, the optimum Step Size is 10.</li>
	</ol>
	</li>
	<li>Click on the Calibration Tool, the icon with the blue ruler, to the right of the Clip Settings button. From New, select Calibration Stick.</li>
	<li>To zoom in on the ruler in the video, right-click on the area to magnify and Select Zoom in from the list. Once appropriately magnified, define the beginning and end of 1 mm on the ruler taped to the microchannel, and Type 1 mm to define the distance.</li>
	<li>Click on the Coordinate Axis Tool, which is the purple icon, to the right of the Calibration Tool. Set the Origin for the x- and y-axis, using the starting frame while doing this step.</li>
	<li>To define the initial point of analysis, create a Point Mass. Click on Create, then select Point Mass. Use Shift + Control to change the size of the rectangle. The initial point is where the inlet and the channel connect. 
	NOTE: The rectangle indicates the domain, defined by the user, that the software will scan to find the contrasting wet and dry powder. The boundary allows the user to define the region where the initial point will be observed.
	<ol>
		<li>Click on Search Next a couple of times to verify that the software is analyzing the correct area. If the software is functioning properly, click on Search and wait for the software to finish analyzing the video. If the software cannot automatically find a matching image intensity from the previous frame to the current frame, the software will stop and wait for the user to redefine the search area. 
		NOTE: For reproducibility and the ability to compare different experimental results, choose the fastest or the slowest point of the fluidic flow front (region of contrast between the wetted and dry powder) for every sample.</li>
		<li>If an analysis error is observed on the live plotted data on the right-hand side of the Tracker screen, click on the Data Point once on the step prior to the erred data point. On the main screen, modify the red rectangular search area location to search the region of interest and repeat step 6.8.1. 
		NOTE: If an error exists, right-click on the inaccurate data point and deselect the point for further analysis.</li>
	</ol>
	</li>
	<li>Once the analysis is complete, copy and paste the results into a spreadsheet. The results saved in the spreadsheet comprise the distance and time data.</li>
	<li>Plot the copied data in the spreadsheet as the distance of fluid transport through the powder bed as a function of time.</li>
</ol>

<ol><li>Redwood, B., Schoffer, F., Garret, B. <a target="_blank" href="http://scholar.google.com/scholar?hl=en&safe=off&q=author%3aRedwood%2C+B+%22The+3D+Printing+Handbook%22">The 3D Printing Handbook</a>. , 3D HUBS. Amsterdam, NL. (2018).</li>
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<li>Ochiai, N., Kraft, E. L., Selker, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Methods+for+colloid+transport+visualization+in+pore+networks%5BTitle%5D)+AND+%22Water+Resources+Research%22%5BJournal%5D)">Methods for colloid transport visualization in pore networks.</a> Water Resources Research. 42 (12), (2006).</li>
<li>Rottman, J., Sierra-Alvarez, R., Shadman, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Real-time+monitoring+of+nanoparticle+retention+in+porous+media%5BTitle%5D)+AND+%22Environmental+Chemistry+Letters%22%5BJournal%5D)">Real-time monitoring of nanoparticle retention in porous media.</a> Environmental Chemistry Letters. 11 (1), 71-76 (2013).</li>
<li>Xing, Y., Chen, X., Chen, X., Zhuang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Colloid-mediated+transport+of+pharmaceutical+and+personal+care+products+through+porous+media%5BTitle%5D)+AND+%22Scientific+Reports%22%5BJournal%5D)">Colloid-mediated transport of pharmaceutical and personal care products through porous media.</a> Scientific Reports. 6 (1), 1-10 (2016).</li>
<li>Dathe, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Functional+models+for+colloid+retention+in+porous+media+at+the+triple+line%5BTitle%5D)+AND+%22Environmental+Science+and+Pollution+Research%22%5BJournal%5D)">Functional models for colloid retention in porous media at the triple line.</a> Environmental Science and Pollution Research. 21 (15), 9067-9080 (2014).</li>
<li>Zhang, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Investigation+of+nanoparticle+adsorption+during+transport+in+porous+media%5BTitle%5D)+AND+%22SPE+Journal%22%5BJournal%5D)">Investigation of nanoparticle adsorption during transport in porous media.</a> SPE Journal. 20 (4), 667-677 (2015).</li>
<li>Zhang, Q., Karadimitriou, N. K., Hassanizadeh, S. M., Kleingeld, P. J., Imhof, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Study+of+colloids+transport+during+two-phase+flow+using+a+novel+polydimethylsiloxane+micro-model%5BTitle%5D)+AND+%22Journal+of+Colloid+and+Interface+Science%22%5BJournal%5D)">Study of colloids transport during two-phase flow using a novel polydimethylsiloxane micro-model.</a> Journal of Colloid and Interface Science. 401, 141-147 (2013).</li>
<li>Environmental Protection Agency. Health and environmental effects of particulate matter (PM). EPA. , Available from: 
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<li>Bridge, J. W., Banwart, S. A., Heathwaite, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Noninvasive+quantitative+measurement+of+colloid+transport+in+mesoscale+porous+media+using+time+lapse+fluorescence+imaging%5BTitle%5D)+AND+%22Environmental+Science+%26+Technology%22%5BJournal%5D)">Noninvasive quantitative measurement of colloid transport in mesoscale porous media using time lapse fluorescence imaging.</a> Environmental Science & Technology. 40 (19), 5930-5936 (2006).</li>
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</ol>

The authors have nothing to disclose.

The protocol that is provided is highly dependent on the material characteristics of the particles that are chosen. Material properties impacting flow include particle size distribution2,3,4,5,11,21, particle surface roughness11, chemical properties at the particle surface2...

Inkjet-based binder jetting into powder media represents an important technology in additive manufacturing (3D printing). The binder jetting process begins with the deposition of functional fluids into powder media using a scanning inkjet printing process. Specifically, an inkjet print head translates over the powder surface, depositing the liquid binding agent onto a powder surface, and thereby forming a solid part in a layer-by-layer fashion1. Inkjet-based binder jetting technologies generally include sand, metal powders, and polymeric powders. However, to expand the materials&#39; space in binder jetting, a fundamental approach to investigating fluid-powder and powder-powder interactions, tribology, powder packing density, and particle aggregation is required. Specifically, for fluid-powder interactions, a critical need exists for the ability to image fluid flow through powder beds in real-time. This promises to be a powerful tool for researchers to include as a characterization technique and potentially as a screening method for different combinations of fluids and powders2,3,4, as well as more complex systems, such as concrete 3D-printing systems that utilize particle-bed methods.
The development of novel imaging techniques of molecular and colloidal transport, including nanoparticles, is an active area of investigation in microfluidic and millifluidic studies. Probing intermolecular interactions by imaging techniques can be challenging, as little work has been done to probe these types of interactions under the conditions of unsaturated and unsteady fluid flow. Many of the studies that are reported in the literature have focused on a saturated, pre-wetted, porous media, such as glass bead5,6,7,8,9,10,11,12 and soils13,14,15,16,17,18. This technique provides a non-invasive approach, resulting in high temporal and spatial resolution2,3,4,19. Furthermore, the developed technique provides a novel method for characterizing and quantifying nano-scale and micron-scale particle transport in a variety of porous media, focusing on polymeric powders.
The proposed technique utilizes a mobile device to record unsaturated, unsteady fluidic transport through porous polymeric media with particle dimensions that are representative of the powders used in 3D printing systems that utilize fluidic powder-bed fusion technologies. This technique is advantageous as the flow cells are cost-effective, reusable, small, and easily handled, illustrating the dominant aspects of frugal science. The ability to implement these simple experiments into a field study is very straightforward, eliminating the complications, cost, and time that are required in optical microscopy. Given the ease of creating the setup, the access to quick results, and the minimal number of sample requirements, this technique is an optimal platform for diagnostic screening.

<table><tbody><tr><td>&amp;micro;-Slide I Luer</td><td>ibidi</td><td>80191</td><td>Microfluidic flow cell</td></tr><tr><td>Beaker</td><td>Southern Labware</td><td>BG1000-800</td><td>Glassware</td></tr><tr><td>CALIBRE 301-58 LT Natural Polycarbonate Resin</td><td>TRINSEO LLC</td><td>CALIBRETM 301-58 LT</td><td>Natural polycarbonate resin</td></tr><tr><td>Ethanol</td><td>Sigma Aldrich</td><td>1.00983</td><td>Solvent</td></tr><tr><td>Fume Hood</td><td>Kewaunee</td><td>Supreme Air LV Fume Hoods</td><td>Used with 92 FPM at 18&quot; opening</td></tr><tr><td>iPhone 7 plus</td><td>Apple</td><td></td><td>Camera</td></tr><tr><td>Opaque 3D printed material</td><td></td><td></td><td>The CAD drawing is provided in the supplemental file</td></tr><tr><td>ORGASOL&amp;nbsp; 2002 ES 6 NAT 3</td><td>ARKEMA</td><td>A12135</td><td>Polyamide powder</td></tr><tr><td>Pipet</td><td>VWR</td><td>10754-268</td><td>Disposable Transfer Pipet</td></tr><tr><td>Pipette</td><td>Globe Scientific Inc.</td><td>3301-200</td><td>Pipette that can hold 125 &amp;micro;L of fluid</td></tr><tr><td>Polystyrene</td><td>Advanced Laser Materials, LLC.</td><td>PS200</td><td>Polystyrene for sintering</td></tr><tr><td>Tracker</td><td></td><td></td><td>Video analysis and modeling tool</td></tr><tr><td>VariQuest 100 White Light Model 3-3700</td><td>FOTODYNE</td><td>&amp;nbsp;3-3700</td><td>White light</td></tr><tr><td>Water</td><td></td><td></td><td>Distilled water</td></tr></tbody></table>

frugal imaging technique of capillary flow through three-dimensional polymeric printing powders

The development of novel imaging techniques of molecular and colloidal transport, including nanoparticles, is an area of active investigation in microfluidic and millifluidic studies. With the advent of three-dimensional (3D) printing, a new domain of materials has emerged, thereby increasing the demand for novel polymers. Specifically, polymeric powders, with average particle sizes on the order of a micron, are experiencing a growing interest from academic and industrial communities. Controlling material tunability at the mesoscopic to microscopic length scales creates opportunities to develop innovative materials, such as gradient materials. Recently, a need for micron-sized polymeric powders has been growing, as clear applications for the material are developing. Three-dimensional printing provides a high-throughput process with a direct link to new applications, driving investigations into the physio-chemical and transport interactions on a mesoscale. The protocol that is discussed in this article provides a non-invasive technique to image fluid flow in packed powder beds, providing high temporal and spatial resolution while leveraging mobile technology that is readily available from mobile devices, such as smartphones. By utilizing a common mobile device, the imaging costs that would normally be associated with an optical microscope are eliminated, resulting in a frugal-science approach. The proposed protocol has successfully characterized a variety of combinations of fluids and powders, creating a diagnostic platform for quickly imaging and identifying an optimal combination of fluid and powder.

In the section on analyzing data, the data for the time-lapsed images in Figure 3 illustrate the 75 wt% ethanol solution infiltrating the polycarbonate (PC) powder. Fluorescein was added to the solution to enhance the image quality for this publication. In the time-lapse images, the time-resolved process begins when the fluid is added to the inlet. Time, t, starts as soon as the fluid begins to penetrate the channel. The series of images demonstrates the progression of the fluid and...

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Frugal Imaging Technique of Capillary Flow Through Three-Dimensional Polymeric Printing Powders

The proposed technique will provide a novel, efficient, frugal, and non-invasive approach for imaging fluidic flow through a packed powder bed, yielding high spatial and temporal resolution.

The development of novel imaging techniques of molecular and colloidal transport, including nanoparticles, is an area of active investigation in ...

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1.2K Views.  South Dakota School of Mines & Technology. The proposed technique will provide a novel, efficient, frugal, and non-invasive approach for imaging fluidic flow through a packed powder bed, yielding high spatial and temporal resolution.

Video: Frugal Imaging Technique of Capillary Flow Through Three-Dimensional Polymeric Printing Powders

Planar and Three-Dimensional Printing of Conductive Inks

Printed electronics rely on low-cost, large-area fabrication routes to create flexible or multidimensional electronic, optoelectronic, and biomedical devices1-3. In this paper, we focus on one- (1D), two- (2D), and three-dimensional (3D) printing of conductive metallic inks in the form of flexible, stretchable, and spanning microelectrodes.
Direct-write assembly4,5 is a 1-to-3D printing technique that enables the fabrication of features ranging from simple lines to complex structures by the deposition of concentrated inks through fine nozzles (~0.1 - 250 &mu;m). This printing method consists of a computer-controlled 3-axis translation stage, an ink reservoir and nozzle, and 10x telescopic lens for visualization. Unlike inkjet printing, a droplet-based process, direct-write assembly involves the extrusion of ink filaments either in- or out-of-plane. The printed filaments typically conform to the nozzle size. Hence, microscale features (&lt; 1 &mu;m) can be patterned and assembled into larger arrays and multidimensional architectures.
In this paper, we first synthesize a highly concentrated silver nanoparticle ink for planar and 3D printing via direct-write assembly. Next, a standard protocol for printing microelectrodes in multidimensional motifs is demonstrated. Finally, applications of printed microelectrodes for electrically small antennas, solar cells, and light-emitting diodes are highlighted.

Planar and three-dimensional printing of conductive metallic inks is described. Our approach provides new avenues for fabricating printed electronic, optoelectronic, and biomedical devices in unusual layouts at the microscale.

Printed electronics rely on low-cost, large-area fabrication routes to create flexible or multidimensional electronic, optoelectronic, and biomedical ...

Bioengineering

Manufacturing of Three-dimensionally Microstructured Nanocomposites through Microfluidic Infiltration

Microstructured composite beams reinforced with complex three-dimensionally (3D) patterned nanocomposite microfilaments are fabricated via nanocomposite in&#64257;ltration of 3D interconnected microfluidic networks. The manufacturing of the reinforced beams begins with the fabrication of microfluidic networks, which involves layer-by-layer deposition of fugitive ink filaments using a dispensing robot, filling the empty space between filaments using a low viscosity resin, curing the resin and finally removing the ink.&#160;Self-supported 3D structures with other geometries and many layers (e.g.&#160;a few hundreds layers) could be built using this method. The resulting tubular micro&#64258;uidic networks are then infiltrated with thermosetting nanocomposite suspensions containing nanofillers (e.g.&#160;single-walled carbon nanotubes), and subsequently cured. The infiltration is done by applying a pressure gradient between two ends of the empty network (either by applying a vacuum or vacuum-assisted microinjection). Prior to the infiltration, the nanocomposite suspensions are prepared by dispersing nanofillers into polymer matrices using ultrasonication and three-roll mixing methods. The nanocomposites (i.e.&#160;materials infiltrated) are then solidified under UV exposure/heat cure, resulting in a 3D-reinforced composite structure. The technique presented here enables the design of functional nanocomposite macroscopic products for microengineering applications such as actuators and sensors.

Three-dimensional (3D) microstructured composite beams are fabricated through the directed and localized infiltration of nanocomposites into 3D porous microfluidic networks. The flexibility of this manufacturing method enables the utilization of different thermosetting materials and nanofillers in order to achieve a variety of functional 3D reinforced nanocomposite macroscopic products.

Microstructured composite beams reinforced with complex three-dimensionally (3D) patterned nanocomposite microfilaments are fabricated via nanocomposite ...

Chemistry

The Submerged Printing of Cells onto a Modified Surface Using a Continuous Flow Microspotter

The printing of cells for microarray applications possesses significant challenges including the problem of maintaining physiologically relevant cell phenotype after printing, poor organization and distribution of desired cells, and the inability to deliver drugs and/or nutrients to targeted areas in the array. Our 3D microfluidic printing technology is uniquely capable of sealing and printing arrays of cells onto submerged surfaces in an automated and multiplexed manner. The design of the microfluidic cell array (MFCA) 3D fluidics enables the printhead tip to be lowered into a liquid-filled well or dish and compressed against a surface to form a seal. The soft silicone tip of the printhead behaves like a gasket and is able to form a reversible seal by applying pressure or backing away. Other cells printing technologies such as pin or ink-jet printers are unable to print in submerged applications. Submerged surface printing is essential to maintain phenotypes of cells and to monitor these cells on a surface without disturbing the material surface characteristics. By printing onto submerged surfaces, cell microarrays are produced that allow for drug screening and cytotoxicity assessment in a multitude of areas including cancer, diabetes, inflammation, infections, and cardiovascular disease.

This 3D microfluidic printing technology prints arrays of cells onto submerged surfaces. We describe how arrays of cells are delivered microfluidically in 3D flow cells onto submerged surfaces. By printing onto submerged surfaces, cell microarrays were produced that allow for drug screening and cytotoxicity assessment in a multitude of areas.

The printing of cells for microarray applications possesses significant challenges including the problem of maintaining physiologically relevant cell ...

Human Cartilage Tissue Fabrication Using Three-dimensional Inkjet Printing Technology

Bioprinting, which is based on thermal inkjet printing, is one of the most attractive enabling technologies in the field of tissue engineering and regenerative medicine. With digital control&#160;cells, scaffolds, and growth factors can be precisely deposited to the desired two-dimensional (2D) and three-dimensional (3D) locations rapidly. Therefore, this technology is an ideal approach to fabricate tissues mimicking their native anatomic structures. In order to engineer cartilage with native zonal organization, extracellular matrix composition (ECM), and mechanical properties, we developed a bioprinting platform using a commercial inkjet printer with simultaneous photopolymerization capable for 3D cartilage tissue engineering. Human chondrocytes suspended in poly(ethylene glycol) diacrylate (PEGDA) were printed for 3D neocartilage construction via layer-by-layer assembly. The printed cells were fixed at their original deposited positions, supported by the surrounding scaffold in simultaneous photopolymerization. The mechanical properties of the printed tissue were similar to the native cartilage. Compared to conventional tissue fabrication, which requires longer UV exposure, the viability of the printed cells with simultaneous photopolymerization was significantly higher. Printed neocartilage demonstrated excellent glycosaminoglycan (GAG) and collagen type II production, which was consistent with gene expression. Therefore, this platform is ideal for accurate cell distribution and arrangement for anatomic tissue engineering.

The methods described in this paper show how to convert a commercial inkjet printer into a bioprinter with simultaneous UV polymerization. The printer is capable of constructing 3D tissue structure with cells and biomaterials. The study demonstrated here constructed a 3D neocartilage.

Bioprinting, which is based on thermal inkjet printing, is one of the most attractive enabling technologies in the field of tissue engineering and ...

Printing Thermoresponsive Reverse Molds for the Creation of Patterned Two-component Hydrogels for 3D Cell Culture

Bioprinting is an emerging technology that has its origins in the rapid prototyping industry. The different printing processes can be divided into contact bioprinting1-4 (extrusion, dip pen and soft lithography), contactless bioprinting5-7 (laser forward transfer, ink-jet deposition) and laser based techniques such as two photon photopolymerization8. It can be used for many applications such as tissue engineering9-13, biosensor microfabrication14-16 and as a tool to answer basic biological questions such as influences of co-culturing of different cell types17. Unlike common photolithographic or soft-lithographic methods, extrusion bioprinting has the advantage that it does not require a separate mask or stamp. Using CAD software, the design of the structure can quickly be changed and adjusted according to the requirements of the operator. This makes bioprinting more flexible than lithography-based approaches.
Here we demonstrate the printing of a sacrificial mold to create a multi-material 3D structure using an array of pillars within a hydrogel as an example. These pillars could represent hollow structures for a vascular network or the tubes within a nerve guide conduit. The material chosen for the sacrificial mold was poloxamer 407, a thermoresponsive polymer with excellent printing properties which is liquid at 4 &deg;C and a solid above its gelation temperature ~20 &deg;C for 24.5% w/v solutions18. This property allows the poloxamer-based sacrificial mold to be eluted on demand and has advantages over the slow dissolution of a solid material especially for narrow geometries. Poloxamer was printed on microscope glass slides to create the sacrificial mold. Agarose was pipetted into the mold and cooled until gelation. After elution of the poloxamer in ice cold water, the voids in the agarose mold were filled with alginate methacrylate spiked with FITC labeled fibrinogen. The filled voids were then cross-linked with UV and the construct was imaged with an epi-fluorescence microscope.

A bioprinter was used to create patterned hydrogels based on a sacrificial mold. The poloxamer mold was backfilled with a second hydrogel and then eluted, leaving voids which were filled with a third hydrogel. This method uses fast elution and good printability of poloxamer to generate complex architectures from biopolymers.

Bioprinting is an emerging technology that has its origins in the rapid prototyping industry. The different printing processes can be divided into contact ...

Immunology and Infection

Multimodal 3D Printing of Phantoms to Simulate Biological Tissue

Biomedical optical imaging is playing an important role in diagnosis and treatment of various diseases. However, the accuracy and the reproducibility of an optical imaging device are greatly affected by the performance characteristics of its components, the test environment, and the operations. Therefore, it is necessary to calibrate these devices by traceable phantom standards. However, most of the currently available phantoms are homogeneous phantoms that cannot simulate multimodal and dynamic characteristics of biological tissue. Here, we show the fabrication of heterogeneous tissue-simulating phantoms using a production line integrating a spin coating module, a polyjet module, a fused deposition modeling (FDM) module, and an automatic control framework. The structural information and the optical parameters of a &#34;digital optical phantom&#34; are defined in a prototype file, imported to the production line, and fabricated layer-by-layer with sequential switch between different printing modalities. Technical capability of such a production line is exemplified by the automatic printing of skin-simulating phantoms that comprise the epidermis, dermis, subcutaneous tissue, and an embedded tumor.

Spin coating, polyjet printing, and fused deposition modeling are integrated to produce multilayered heterogeneous phantoms that simulate structural and functional properties of biological tissue.

Biomedical optical imaging is playing an important role in diagnosis and treatment of various diseases. However, the accuracy and the reproducibility of ...

Triplet Fusion Upconversion Nanocapsule Synthesis

Triplet fusion upconversion (UC) allows for the generation of one high energy photon from two low energy input photons. This well-studied process has significant implications for producing high energy light beyond a material's surface. However, the deployment of UC materials has been stymied due to poor material solubility, high concentration requirements, and oxygen sensitivity, ultimately resulting in reduced light output. Toward this end, nanoencapsulation has been a popular motif to circumvent these challenges, but durability has remained elusive in organic solvents. Recently, a nanoencapsulation technique was engineered to tackle each of these challenges, whereupon an oleic acid nanodroplet containing upconversion materials was encapsulated with a silica shell. Ultimately, these nanocapsules (NCs) were durable enough to enable triplet fusion upconversion-facilitated volumetric three-dimensional (3D) printing. By encapsulating upconversion materials with silica and dispersing them in a 3D printing resin, photopatterning beyond the surface of the printing vat was made possible. Here, video protocols for the synthesis of upconversion NCs are presented for both small-scale and large-scale batches. The outlined protocols serve as a starting point for adapting this encapsulation scheme to multiple upconversion schemes for use in volumetric 3D printing applications.

This protocol details the synthesis of upconversion nanocapsules for subsequent use in photopolymerizable resins for triplet fusion upconversion-facilitated volumetric 3D printing.

Triplet fusion upconversion (UC) allows for the generation of one high energy photon from two low energy input photons. This well-studied process has ...

Visualizing Polymer Dynamics for Optimizing 3D Printing Quality

Real-Time Imaging of Bonding in 3D-Printed Layers

In recent times, 3D printing technology has revolutionized our ability to design and produce products, but optimizing the print quality can be challenging. The process of extrusion 3D printing involves pressuring molten material through a thin nozzle and depositing it onto previously extruded material. This method relies on bonding between the consecutive layers to create a strong and visually appealing final product. This is no easy task, as many parameters, such as the nozzle temperature, layer thickness, and printing speed, must be fine-tuned to achieve optimal results. In this study, a method for visualizing the polymer dynamics during extrusion is presented, giving insight into the layer bonding process. Using laser speckle imaging, the plastic flow and fusion can be resolved non-invasively, internally, and with high spatiotemporal resolution. This measurement, which is easy to perform, provides an in-depth understanding of the underlying mechanics influencing the final print quality. This methodology was tested with a range of cooling fan speeds, and the results showed increased polymer motion with lower fan speeds and, thus, explained the poor printing quality when the cooling fan was turned off. These findings show that this methodology allows for optimizing the printing settings and understanding the material behavior. This information can be used for the development and testing of novel printing materials or advanced slicing procedures. With this approach, a deeper understanding of extrusion can be built to take 3D printing to the next level.

Author Spotlight: Real-Time Imaging of Bonding in 3D-Printed Layers  

With a non-invasive and real-time technique, nanoscopic polymer motion inside a polymer filament is imaged during 3D printing. Fine-tuning this motion is crucial for producing constructs with optimal performance and appearance. This method reaches the core of plastic layer fusion, thus offering insights into optimal printing conditions and material design criteria.

In recent times, 3D printing technology has revolutionized our ability to design and produce products, but optimizing the print quality can be ...

Three-dimensional Printing of Thermoplastic Materials to Create Automated Syringe Pumps with Feedback Control for Microfluidic Applications

Microfluidics has become a critical tool in research across the biological, chemical, and physical sciences. One important component of microfluidic experimentation is a stable fluid handling system capable of accurately providing an inlet flow rate or inlet pressure. Here, we have developed a syringe pump system capable of controlling and regulating the inlet fluid pressure delivered to a microfluidic device. This system was designed using low-cost materials and additive manufacturing principles, leveraging three-dimensional (3D) printing of thermoplastic materials and off-the-shelf components whenever possible. This system is composed of three main components: a syringe pump, a pressure transducer, and a programmable microcontroller. Within this paper, we detail a set of protocols for fabricating, assembling, and programming this syringe pump system. Furthermore, we have included representative results that demonstrate high-fidelity, feedback control of inlet pressure using this system. We expect this protocol will allow researchers to fabricate low-cost syringe pump systems, lowering the entry barrier for the use of microfluidics in biomedical, chemical, and materials research.

Here we present a protocol to construct a pressure-controlled syringe pump to be used in microfluidic applications. This syringe pump is made from an&#160;additively manufactured body, off-the-shelf hardware, and open-source electronics. The resulting system is low-cost, straightforward to build, and delivers well-regulated fluid flow to enable rapid microfluidic research.

Microfluidics has become a critical tool in research across the biological, chemical, and physical sciences. One important component of microfluidic ...

3D Printing of In Vitro Hydrogel Microcarriers by Alternating Viscous-Inertial Force Jetting

Microcarriers are beads with a diameter of 60-250 &#181;m and a large specific surface area, which are commonly used as carriers for large-scale cell cultures. Microcarrier culture technology has become one of the main techniques in cytological research and is commonly used in the field of large-scale cell expansion. Microcarriers have also been shown to play an increasingly important role in in vitro tissue engineering construction and clinical drug screening. Current methods for preparing microcarriers include microfluidic chips and inkjet printing, which often rely on complex flow channel design, an incompatible two-phase interface, and a fixed nozzle shape. These methods face the challenges of complex nozzle processing, inconvenient nozzle changes, and excessive extrusion forces when applied to multiple bioink. In this study, a 3D printing technique, called alternating viscous-inertial force jetting, was applied to enable the construction of hydrogel microcarriers with a diameter of 100-300 &#181;m. Cells were subsequently seeded on microcarriers to form tissue engineering modules. Compared to existing methods, this method offers a free nozzle tip diameter, flexible nozzle switching, free control of printing parameters, and mild printing conditions for a wide range of bioactive materials.

3D Printing of In Vitro Hydrogel Microcarriers by Alternating Viscous-Inertial Force Jetting

Presented here is a mild 3D printing technique driven by alternating viscous-inertial forces to enable the construction of hydrogel microcarriers. Homemade nozzles offer flexibility, allowing easy replacement for different materials and diameters. Cell binding microcarriers with a diameter of 50-500 &#181;m can be obtained and collected for further culturing.

Microcarriers are beads with a diameter of 60-250 &#181;m and a large specific surface area, which are commonly used as carriers for large-scale cell ...

Frugal Imaging Technique of Capillary Flow Through Three-Dimensional Polymeric Printing Powders

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