This work was sponsored by the National Natural Science Foundation of China (51375176); the Guangdong Provincial Natural Science Foundation of China (2014A030313264); and the Science and Technology Planning Project of Guangdong Province, China (2014B010126003).

Caution: Please consult all relevant material safety data sheets (MSDS) before use. Acetone is highly flammable and can cause irritation to the eyes and respiratory tract. The voltage involved is as high as several thousand volts; hence, electrical sparks are expected when conducting the experiment. Carry out the experiments in a room with good ventilation to avoid explosions and fire from the sparks.
1. Fabrication of the Plates and Holder
NOTE: In this work, the electrode plates and holder are fabricated by a production line in a factory. Only the material and the parameters of all parts in this paper will be introduced due to the complicated processes.
<ol>
	<li>Material and size of the electrode plate
	<ol>
		<li>Fabricate the electrode plates using 1.4 mm FR-4 CCL with a thin copper layer of 35 &#181;m. See Figure 1 for detailed parameters of the electrode plate.</li>
	</ol>
	</li>
	<li>Parameters of the electrodes
	<ol>
		<li>Order the electrode plates from the factory. See Figure 2 for more details.</li>
	</ol>
	</li>
	<li>Inspection of the electrode plate
	<ol>
		<li>After the preparation of the electrode plate, use an electron microscope to inspect the electrodes for any noticeable flaws under 100X and 300X magnification. Note that any tiny defects on the surface of the electrodes can cause short-circuiting, as shown in Figure 3.</li>
		<li>Inspect and measure the electrode width and spacing to determine whether the dimension accuracy meets the requirement.</li>
		<li>Test the plate with an amperemeter to see if an electrical short-circuit occurs.</li>
	</ol>
	</li>
	<li>Preparation of the chamber plate
	<ol>
		<li>Cut some silicone membrane to the same size as the electrode plate, as shown in Figure 4. Choose silicone membranes with different thicknesses to make chamber plates with different heights.</li>
		<li>Use a special punching tool to punch the chamber hole, as shown in Figure 5.</li>
	</ol>
	</li>
	<li>Processing of the holder
	<ol>
		<li>Order the holder from a factory. The detailed parameters are shown in Figure 6.</li>
	</ol>
	</li>
	<li>Fabrication of the cover plate
	<ol>
		<li>Drill two holes on the top of the cover plate using a drilling machine to install the inlet and outlet tubes. See Figure 7 for their positions and sizes.</li>
	</ol>
	</li>
</ol>
2. Assembly of the Micropump
<ol>
	<li>Use acetone to wash all the plates, the holder, the inlet and outlet tubes, and other tools used in the experiments. Put these tools and plates inside a beaker and then pour enough 99.5% acetone to immerse them. Put the beaker inside the ultrasonic washer. Turn on the ultrasonic washer and set the timer to 5 min.</li>
	<li>Insert the inlet and outlet stainless-steel tubes into the two holes on the cover plate.</li>
	<li>Place a chamber plate made of silicone membrane on the electrode plate and then cover it with the cover plate.</li>
	<li>Stack and align the cover plate, the chamber plate, and the electrode plate from top to bottom and insert the aligned plates into the holder.
	<ol>
		<li>Use an M5 bolt to fix the plates inside the holder. See the explosion view and normal view of the assembled micropump, as shown in Figure 8 and Figure 9, respectively.</li>
		<li>Press the plates together by tightening the bolts. 
		NOTE: The tubes and the cavity on the chamber plate will form a passage for the working liquid. The elastic chamber plate can also seal off the gap between plates to prevent liquid from flowing out. See the explosion view and the normal view of the assembled micropump in Figure 8 and Figure 9, respectively.</li>
	</ol>
	</li>
	<li>Use two polyurethane hoses with external diameters of 4 mm and internal diameters of 2 mm to connect the inlet and outlet stainless-steel tubes.</li>
	<li>Connect an amperemeter, a 500 V DC power source, and the micropump in series. Insert a 1 mA fuse between the amperemeter and the power source to protect the amperemeter in case the micropump is shorted.</li>
	<li>Insert the inlet hose into a 50 mL beaker with 20-30 mL of acetone inside. 
	NOTE: Figure 10 shows the completed platform.</li>
</ol>
3. Experimental Procedure
<ol>
	<li>Preparatory work before the experiment
	<ol>
		<li>Use a cylinder to inject acetone to fill up the micropump. After the liquid level reaches the outlet hose, continue to inject 10 mL of acetone inside until all bubbles are pushed away from the chamber. 
		NOTE: It is impossible to see if there are any bubbles left inside the chamber because the cover plate and the electrode plate are not transparent. Continuously injecting acetone helps to remove bubbles, but it cannot guarantee that no bubbles are left inside the micropump. Bubbles may block the passage of liquid, or they may short the circuits and cause a micro-explosion inside the micropump, which will burn the electrodes. The effect of bubbles on the pump operation is not totally clear yet, but the breakdowns they cause have been observed several times.</li>
		<li>Pour 20-30 mL of acetone into the beaker and put the inlet hose inside the beaker. Ensure that the liquid level is at least 5 mm higher than the inlet so that acetone can flow into the pump and no air can be sucked into the micropump chamber.</li>
	</ol>
	</li>
	<li>Static pressure test
	<ol>
		<li>Attach the outlet hose to a small frame so that the hose can remain straight and vertical. Put a ruler alongside the outlet hose to measure the liquid level.</li>
		<li>Connect the micropump to the power source.</li>
		<li>Start the test by pressing the switch and then mark down the initial liquid level.</li>
		<li>After the liquid level becomes stable, record the time, the final liquid level, and the electric current.</li>
		<li>Continue to record the liquid level and the current every 10 s until the micropump breaks down.</li>
	</ol>
	</li>
	<li>Flow rate test
	<ol>
		<li>Use a large measuring cylinder to collect the liquid coming out of the outlet hose. Be sure to fix the outlet hose so that the end remains at the same altitude as the liquid level in the beaker.</li>
		<li>Connect the micropump to the power source.</li>
		<li>Start the test by pressing the switch and then mark down the initial liquid level.</li>
		<li>As the liquid starts to flow out of the outlet hose, record the volume of acetone inside the measuring cylinder every 10 s. As the experiment goes on, add acetone to the beaker to maintain the liquid level.</li>
	</ol>
	</li>
	<li>Reliability test
	<ol>
		<li>Use the average working time to evaluate the reliability of the pump. During the flow rate test and the static pressure test, record the operation time before the pump breaks down. Record the detailed phenomena of each breakdown during the experiment and inspect the electrode plate surface afterwards for further analysis.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>Kazemi, P. Z., Selvaganapathy, P. R., Ching, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+micropillar+electrode+spacing+on+the+performance+of+electrohydrodynamic+micropumps.">Effect of micropillar electrode spacing on the performance of electrohydrodynamic micropumps.</a> J Electrostat. 68 (4), 376-383 (2010).</li><li>Kano, I., Nishina, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+electrode+arrangements+on+EHD+conduction+pumping.">Effect of electrode arrangements on EHD conduction pumping.</a> IEEE Trans Ind Appl. 49 (2), 679-684 (2013).</li><li>Laser, D. J., Santiago, J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+of+micropumps.">A review of micropumps.</a> J Micromech Microeng. 14 (6), R35 (2004).</li><li>Fylladitakis, E. D., Theodoridis, M. P., Moronis, A. X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Review+on+the+history,+research,+and+applications+of+electrohydrodynamics.">Review on the history, research, and applications of electrohydrodynamics.</a> IEEE Trans Plasma Sci. 42 (2), 358-375 (2014).</li><li>Yazdani, M., Seyed-Yagoobi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Electrostatics Joint Conf. , (2009).</li><li>Gharraei, R., Esmaeilzadeh, E., Hemayatkhah, M., Danaeefar, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+investigation+of+electrohydrodynamic+conduction+pumping+of+various+liquids+film+using+flush+electrodes.">Experimental investigation of electrohydrodynamic conduction pumping of various liquids film using flush electrodes.</a> J Electrostat. 69 (1), 43-53 (2011).</li><li>Gharraei, R., Esmaeilzadeh, E., Nobari, M. R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Numerical+investigation+of+conduction+pumping+of+dielectric+liquid+film+using+flush-mounted+electrodes.">Numerical investigation of conduction pumping of dielectric liquid film using flush-mounted electrodes.</a> Theor Comp Fluid Dyn. 28 (1), 89 (2014).</li><li>Jeong, S. -. I., Seyed-Yagoobi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+study+of+electrohydrodynamic+pumping+through+conduction+phenomenon.">Experimental study of electrohydrodynamic pumping through conduction phenomenon.</a> J Electrostat. 56 (2), 123-133 (2002).</li><li>Seyed-Yagoobi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrohydrodynamic+pumping+of+dielectric+liquids.">Electrohydrodynamic pumping of dielectric liquids.</a> J Electrostat. 63 (6), 861-869 (2005).</li><li>Hojjati, M., Esmaeilzadeh, E., Sadri, B., Gharraei, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrohydrodynamic+conduction+pumps+with+cylindrical+electrodes+for+pumping+of+dielectric+liquid+film+in+an+open+channel.">Electrohydrodynamic conduction pumps with cylindrical electrodes for pumping of dielectric liquid film in an open channel.</a> Colloid Surface A. 392 (1), 294-299 (2011).</li><li>Yazdani, M., Seyed-Yagoobi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Numerical+investigation+of+electrohydrodynamic-conduction+pumping+of+liquid+film+in+the+presence+of+evaporation.">Numerical investigation of electrohydrodynamic-conduction pumping of liquid film in the presence of evaporation.</a> J Heat Trans-T ASME. 131 (1), 011602 (2009).</li><li>Vafaie, R. H., Ghavifekr, H. B., Lintel, H., Brugger, J., Renaud, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bi-directional+AC+electrothermal+micropump+for+on-chip+biological+applications.">Bi-directional AC electrothermal micropump for on-chip biological applications.</a> Electrophoresis. 37 (5-6), 719-726 (2016).</li><li>Pearson, M. R., Seyed-Yagoobi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+Study+of+Linear+and+Radial+Two-Phase+Heat+Transport+Devices+Driven+by+Electrohydrodynamic+Conduction+Pumping.">Experimental Study of Linear and Radial Two-Phase Heat Transport Devices Driven by Electrohydrodynamic Conduction Pumping.</a> J Heat Trans-T ASME. 137 (2), 022901 (2015).</li></ol>

The authors have nothing to disclose.

One of the critical steps within the protocol is to inspect the electrode plate carefully. Small burrs on the edge of an electrode can result in a short-circuit, and surface integrity can greatly affect pump performance. The cleaning of the electrode plate and holder is also very important. The electrode chamber height is less than 1 mm, so small dust particles may block the working liquid flow and cause a short-circuit. Before the test, injecting acetone into the chamber can remove the bubbles outside of the chamber.
The performance of the micropump can be severely influenced by the height of chamber. To eliminate such influence, the elastic chamber plate can be replaced by harder material.
There are a few limitations to the technique. First, the electrode height is determined by the metal layer on the FR-4 CCL. It is relatively high, which affects the flow of liquid to some extent. Second, by using silicone membrane as the chamber plate, the sealability of the chamber is improved. However, the elasticity of silicone can cause some deviation in chamber height. Finally, the electrode width and electrode spacing are limited by the technique itself, which makes achieving higher electric field strength much more difficult than with other technique.
Unlike photolithography and other high-precision fabrication techniques, using FR-4 CCL with this electrode fabrication process is relatively simpler and cheaper. On the other hand, the required electric voltage in this work is much lower. Pearson and Seyed-Yagoobi13 have proposed a ring electrode and perforated electrode design that requires 5 kV of DC voltage for the pump to work, while in this work, the power source is only 500 V.
This micropump can be used to loop a heat pipe, especially a long heat pipe that requires a driving force other than capillary force. Inserting one or multiple electrode plates inside the heat pipe can provide enough driving force for the dielectric coolant to travel from the cold end to the hot end. A similar function can also be achieved by an annular electrode fabricated on soft, insulated material with a metal substrate.

Micropumps can drive liquid flow on a much smaller scale than most pumps. In recent years, various driving schemes have been applied successfully to microfluidic systems1,2,3,4,5. The electrohydrodynamic (EHD) pump can exert forces directly on the liquid, without any moving parts, which makes it simpler and easier to fabricate6. According to the charge types, EHD pumps can be classified as injection pumps, induction pumps, or conduction pumps. Induction pumps do not work on isothermal liquids, while injection pumps change the liquid conductivity. Because they lack such problems, conduction pumps are more stable and have a wider application.
The conduction pump is based on the mismatch of the dissociation and recombination rates of liquid molecules. Normally, the dissociation and recombination process can be expressed as follows7,8: 
<img alt="Equation" src="/files/ftp_upload/55867/55867eq1.jpg" /> 
where the recombination rate kr is constant while the dissociation rate kd is a function of the electric field strength. When the electric field strength reaches a certain value, the dissociation rate will exceed the recombination rate. Then, more and more free charges travel to the two electrodes of opposite polarity, and heterocharge layers form. These heterocharge layers are the key to the pump, as the movement of the charges pushes the liquid molecules forward. Therefore, net body force can be generated in the liquid within the chamber using asymmetric electrodes or the mismatch of the mobility of positive and negative ions9,10,11,12.
This work introduces a new way of fabricating a symmetric planar electrode plate for a conduction pump. The electrode plate is prepared on FR-4 CCL, and the pump chamber is prepared by micromachining. The fabrication processes are relatively simpler and more convenient than those of other manufacturing methods, such as nanolithography. A testing platform is set up to investigate the performance of the conduction micropump under different conditions. Furthermore, the reliability of the conduction micropump is also investigated under different circumstances.

<table width="683">
	<tbody>
		<tr height="21">
			<td height="21" style="width:207px;height:21px;">Amperemeter</td>
			<td style="width:311px;">-</td>
			<td style="width:101px;">85C1-MA</td>
			<td style="width:64px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="width:207px;height:21px;">DC high voltage power supply</td>
			<td style="width:311px;">NanTong Jianuo electric device company</td>
			<td style="width:101px;">GY-WY500-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="width:207px;height:21px;">Fuse</td>
			<td style="width:311px;">-</td>
			<td style="width:101px;">-</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="width:207px;height:21px;">Ultrasonic cleaner</td>
			<td style="width:311px;">Derui ultrasonic device company</td>
			<td style="width:101px;">-</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="width:207px;height:21px;">Soldering iron</td>
			<td style="width:311px;">-</td>
			<td style="width:101px;">-</td>
			<td></td>
		</tr>
	</tbody>
</table>

a performance-testing platform for a conduction micropump with an fr-4 copper-clad electrode plate

Here, a conduction micropump with symmetric planar electrode pairs prepared on flame-retardant glass-reinforced epoxy (FR-4) copper-clad laminate (CCL) is fabricated. It is used to investigate the influence of chamber dimensions on the performance of a conduction micropump and to determine the reliability of the conduction pump when acetone is used as the working fluid. A testing platform is set up to evaluate conduction micropump performance under different conditions. When the chamber height is 0.2 mm, the pump pressure reaches its peak value.

As shown in Figure 11, the pump pressure and its increasing rate rise when the voltage increases. When the voltage reaches 500 V, the pump pressure reaches 1,100 Pa.
The pump static pressure rises with the pump chamber height increasing when the chamber height is under 0.2 mm. The pump performance reaches its highest point when the chamber height is 0.2 mm. Then, the static pressure drops when the chamber height continues to increase. It is believed that 0.2 mm is the best value for the chamber height. The results are shown in Figure 12.
By increasing the chamber length, the static pressure rises, with a big slope until a chamber length of 23 mm and a smaller slope, indicating a gradual rise, afterwards. The variation tendency of the pump pressure is shown in Figure 13.
As shown in Figure 14, there is a slight decrease in the static pressure versus chamber width curve when the chamber width increases from 2 mm to 3 mm. Afterwards, the static pressure remains at a certain level as the chamber width increases.
During testing, the micropump breaks down frequently after working for 10-90 min. After working for a certain amount of time, the pump pressure continues to drop until breakdown occurs. However, the performance of the pump pressure can be restored when new acetone is added to the beaker.
When there are bubbles inside the pump, the pump pressure will not rise as high as it normally does because the passage of working liquid is blocked by the bubbles. If the plates are not sufficiently cleaned, meaning that there is still dust or other contamination, the pump will easily get shorted when these particles travel to the gap between electrodes. When the pump gets shorted, the electric current will rise very fast and burn the electrodes.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/55867/55867fig1.jpg" /> 
Figure 1: Size of the electrode plate. The electrode plate, chamber plate, and cover plate are of the same size. Their widths are 9 mm and their lengths are 40 mm. The holes for soldering the wire have a diameter of 1.6 mm. Order the electrode plates from the factory with these parameters and check the plate size afterwards to make sure that they can fit inside the holder. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/55867/55867fig2.jpg" /> 
Figure 2: Dimensions of the electrodes. The electrode plates have 23 pairs of electrodes, with an electrode width of 300 &#181;m, an electrode spacing of 200 &#181;m, a width gap between the two electrode pairs of 400 &#181;m, and an electrode end spacing of 600 &#181;m. The electrode spacing is the most important parameter, as it determines the electric field strength. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/55867/55867fig3.jpg" /> 
Figure 3: Tiny defects on the surface of electrodes and the damage from short circuits. Rags and surface pits are the typical defects of the electrodes. The pits in (a) have caused a severe breakdown. Some of the electrodes in (b) burn under high electric field strength. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/55867/55867fig4.jpg" /> 
Figure 4: Chamber plate dimensions. The width and length of the plate are 9 mm and 40 mm, respectively. The thickness of the plate is 0.3 mm in these tests. The dimension accuracy is 0.1 mm. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/55867/55867fig5.jpg" /> 
Figure 5: Picture of the special hole punch. The special punching tool is customized for cutting the cavity on the chamber plate. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/55867/55867fig6.jpg" /> 
Figure 6: The engineering drawing of the holder. The 3 mm opening on the top is for the inlet and outlet stainless-steel tubes. The thickness of all walls is 2 mm, and the overall size of the holder is 14 x 37.5 x 9 mm3. The 2.4 mm wall on the left is for holding the position of the electrode plate, cover plate, and chamber plate. On the bottom of the holder, there is an M5 threaded hole for a fastening bolt. The dimension accuracy of the holder is 0.1 mm, which is not very high. Make sure that the electrode plate can fit inside. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/55867/55867fig7.jpg" /> 
Figure 7: The parameters of the cover plate. The material of the cover plate is also FR4. Two stainless-steel tubes are inserted into the hole to provide inlet and outlet passage for the working liquid. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/55867/55867fig8.jpg" /> 
Figure 8: Explosion view of the micropump. From top to bottom are the inlet and outlet tubes, the cover plate, the chamber plate, the electrode plate, and the fastening bolt. The tubes and the cavity on the chamber plate form a liquid passage for the working liquid. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig8large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 9" class="xfigimg" src="/files/ftp_upload/55867/55867fig9.jpg" /> 
Figure 9: Rendered image of the assembled micropump. After fastening the bolt, all plates are pressed together, and the hole on the chamber plate is the only space for liquid to flow through. The elastic chamber plate can also seal off the gap between plates to prevent liquid from flowing out. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig9large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 10" class="xfigimg" src="/files/ftp_upload/55867/55867fig10.jpg" /> 
Figure 10: The picture of the micropump experiment platform. The bottle that contains acetone can be replace by a beaker, but it is safer to cover the beaker with thin film to prevent the acetone from volatilizing too much. The power source, the amperemeter, and the pump are connected in series. The outlet hose can be also replaced by a glass tube. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig10large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 11" class="xfigimg" src="/files/ftp_upload/55867/55867fig11v2.jpg" /> 
Figure 11: The relationship between the voltage and the pump pressure. The pump pressure and its increasing rate rises when the voltage increases. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig11v2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 12" class="xfigimg" src="/files/ftp_upload/55867/55867fig12v2.jpg" /> 
Figure 12: The relationship between the chamber height and the pump performance. The pump performance first increases then drops when the chamber height increases from 0.1 mm to 0.5 mm. <a href="https://cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55867/55867fig12v2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 13" class="xfigimg" src="/files/ftp_upload/55867/55867fig13v2.jpg" /> 
Figure 13: The relationship between the chamber length and the pump performance. The pump performance rises when the length increases.
<img alt="Figure 14" class="xfigimg" src="/files/ftp_upload/55867/55867fig14v2.jpg" /> 
Figure 14: The relationship between the chamber width and the pump performance. The pump pressure remains the same when the chamber width increases.

Watch this Scientific Journal Video about A Performance-testing Platform for a Conduction Micropump with an FR-4 Copper-clad Electrode Plate at JoVE.com

A Performance-testing Platform for a Conduction Micropump with an FR-4 Copper-clad Electrode Plate

This paper presents a protocol for the fabrication of a conduction micropump using symmetric planar electrodes on flame-retardant glass-reinforced epoxy (FR-4) copper-clad laminate (CCL) to test the influence of chamber dimensions on the performance of a conduction micropump.

Here, a conduction micropump with symmetric planar electrode pairs prepared on flame-retardant glass-reinforced epoxy (FR-4) copper-clad laminate (CCL) is ...

a-performance-testing-platform-for-conduction-micropump-with-an-fr-4

Nanyang Technological University

Research

JoVE Journal

Bioengineering

5.9K Views.  South China University of Technology. This paper presents a protocol for the fabrication of a conduction micropump using symmetric planar electrodes on flame-retardant glass-reinforced epoxy (FR-4) copper-clad laminate (CCL) to test the influence of chamber dimensions on the performance of a conduction micropump.

Video: A Performance-testing Platform for a Conduction Micropump with an FR-4 Copper-clad Electrode Plate

Environmentally-controlled Microtensile Testing of Mechanically-adaptive Polymer Nanocomposites for ex vivo Characterization

Implantable microdevices are gaining significant attention for several biomedical applications1-4. Such devices have been made from a range of materials, each offering its own advantages and shortcomings5,6. Most prominently, due to the microscale device dimensions, a high modulus is required to facilitate implantation into living tissue. Conversely, the stiffness of the device should match the surrounding tissue to minimize induced local strain7-9. Therefore, we recently developed a new class of bio-inspired materials to meet these requirements by responding to environmental stimuli with a change in mechanical properties10-14. Specifically, our poly(vinyl acetate)-based nanocomposite (PVAc-NC) displays a reduction in stiffness when exposed to water and elevated temperatures (e.g. body temperature). Unfortunately, few methods exist to quantify the stiffness of materials in vivo15, and mechanical testing outside of the physiological environment often requires large samples inappropriate for implantation. Further, stimuli-responsive materials may quickly recover their initial stiffness after explantation. Therefore, we have developed a method by which the mechanical properties of implanted microsamples can be measured ex vivo, with simulated physiological conditions maintained using moisture and temperature control13,16,17. 
To this end, a custom microtensile tester was designed to accommodate microscale samples13,17 with widely-varying Young's moduli (range of 10 MPa to 5 GPa). As our interests are in the application of PVAc-NC as a biologically-adaptable neural probe substrate, a tool capable of mechanical characterization of samples at the microscale was necessary. This tool was adapted to provide humidity and temperature control, which minimized sample drying and cooling17. As a result, the mechanical characteristics of the explanted sample closely reflect those of the sample just prior to explantation.
The overall goal of this method is to quantitatively assess the in vivo mechanical properties, specifically the Young's modulus, of stimuli-responsive, mechanically-adaptive polymer-based materials. This is accomplished by first establishing the environmental conditions that will minimize a change in sample mechanical properties after explantation without contributing to a reduction in stiffness independent of that resulting from implantation. Samples are then prepared for implantation, handling, and testing (Figure 1A). Each sample is implanted into the cerebral cortex of rats, which is represented here as an explanted rat brain, for a specified duration (Figure 1B). At this point, the sample is explanted and immediately loaded into the microtensile tester, and then subjected to tensile testing (Figure 1C). Subsequent data analysis provides insight into the mechanical behavior of these innovative materials in the environment of the cerebral cortex.

Environmentally-controlled Microtensile Testing of Mechanically-adaptive Polymer Nanocomposites for ex vivo Characterization

A method is discussed by which the in vivo mechanical behavior of stimuli-responsive materials is monitored as a function of time. Samples are tested ex vivo using a microtensile tester with environmental controls to simulate the physiological environment. This work further promotes understanding the in vivo behavior of our material.

Implantable microdevices are gaining significant attention for several biomedical applications1-4. Such devices have been made from a range of materials, ...

A Versatile Automated Platform for Micro-scale Cell Stimulation Experiments

Study of cells in culture (in vitro analysis) has provided important insight into complex biological systems. Conventional methods and equipment for in vitro analysis are well suited to study of large numbers of cells (&ge;105) in milliliter-scale volumes (&ge;0.1 ml). However, there are many instances in which it is necessary or desirable to scale down culture size to reduce consumption of the cells of interest and/or reagents required for their culture, stimulation, or processing. Unfortunately, conventional approaches do not support precise and reproducible manipulation of micro-scale cultures, and the microfluidics-based automated systems currently available are too complex and specialized for routine use by most laboratories. To address this problem, we have developed a simple and versatile technology platform for automated culture, stimulation, and recovery of small populations of cells (100 - 2,000 cells) in micro-scale volumes (1 - 20 &mu;l). The platform consists of a set of fibronectin-coated microcapillaries ("cell perfusion chambers"), within which micro-scale cultures are established, maintained, and stimulated; a digital microfluidics (DMF) device outfitted with "transfer" microcapillaries ("central hub"), which routes cells and reagents to and from the perfusion chambers; a high-precision syringe pump, which powers transport of materials between the perfusion chambers and the central hub; and an electronic interface that provides control over transport of materials, which is coordinated and automated via pre-determined scripts. As an example, we used the platform to facilitate study of transcriptional responses elicited in immune cells upon challenge with bacteria. Use of the platform enabled us to reduce consumption of cells and reagents, minimize experiment-to-experiment variability, and re-direct hands-on labor. Given the advantages that it confers, as well as its accessibility and versatility, our platform should find use in a wide variety of laboratories and applications, and prove especially useful in facilitating analysis of cells and stimuli that are available in only limited quantities.

We have developed an automated cell culture and interrogation platform for micro-scale cell stimulation experiments. The platform offers simple, versatile, and precise control in cultivating and stimulating small populations of cells, and recovering lysates for molecular analyses. The platform is well suited to studies that use precious cells and/or reagents.

Study of cells in culture (in vitro analysis) has provided important insight into complex biological systems. Conventional methods and equipment for in ...

Investigating Receptor-ligand Systems of the Cellulosome with AFM-based Single-molecule Force Spectroscopy

Cellulosomes are discrete multienzyme complexes used by a subset of anaerobic bacteria and fungi to digest lignocellulosic substrates. Assembly of the enzymes onto the noncatalytic scaffold protein is directed by interactions among a family of related receptor-ligand pairs comprising interacting cohesin and dockerin modules. The extremely strong binding between cohesin and dockerin modules results in dissociation constants in the low picomolar to nanomolar range, which may hamper accurate off-rate measurements with conventional bulk methods. Single-molecule force spectroscopy (SMFS) with the atomic force microscope measures the response of individual biomolecules to force, and in contrast to other single-molecule manipulation methods (i.e.&#160;optical tweezers), is optimal for studying high-affinity receptor-ligand interactions because of its ability to probe the high-force regime (&#62;120 pN). Here we present our complete protocol for studying cellulosomal protein assemblies at the single-molecule level. Using a protein topology derived from the native cellulosome, we worked with enzyme-dockerin and carbohydrate binding module-cohesin (CBM-cohesin) fusion proteins, each with an accessible free thiol group at an engineered cysteine residue. We present our site-specific surface immobilization protocol, along with our measurement and data analysis procedure for obtaining detailed binding parameters for the high-affinity complex. We demonstrate how to quantify single subdomain unfolding forces, complex rupture forces, kinetic off-rates, and potential widths of the binding well. The successful application of these methods in characterizing the cohesin-dockerin interaction responsible for assembly of multidomain cellulolytic complexes is further described.

Cellulosomes are multienzyme complexes designed for digesting cellulose. AFM-based SMFS was used to study the mechanical properties and folding configuration of cellulosome-associated protein assemblies. We present a complete workflow for protein immobilization, data acquisition, and data analysis to study the interactions of individual receptor-ligand complexes involved in cellulosome assembly.

Cellulosomes are discrete multienzyme complexes used by a subset of anaerobic bacteria and fungi to digest lignocellulosic substrates. Assembly of the ...

Cell Squeezing as a Robust, Microfluidic Intracellular Delivery Platform

Rapid mechanical deformation of cells has emerged as a promising, vector-free method for intracellular delivery of macromolecules and nanomaterials. This technology has shown potential in addressing previously challenging applications; including, delivery to primary immune cells, cell reprogramming, carbon nanotube, and quantum dot delivery. This vector-free microfluidic platform relies on mechanical disruption of the cell membrane to facilitate cytosolic delivery of the target material. Herein, we describe the detailed method of use for these microfluidic devices including, device assembly, cell preparation, and system operation. This delivery approach requires a brief optimization of device type and operating conditions for previously unreported applications. The provided instructions are generalizable to most cell types and delivery materials as this system does not require specialized buffers or chemical modification/conjugation steps. This work also provides recommendations on how to improve device performance and trouble-shoot potential issues related to clogging, low delivery efficiencies, and cell viability.

Rapid mechanical deformation of cells has emerged as a promising, vector-free method for intracellular delivery of macromolecules and nanomaterials. This protocol provides detailed steps on how to use the system for a broad range of applications.

Rapid mechanical deformation of cells has emerged as a promising, vector-free method for intracellular delivery of macromolecules and nanomaterials. This ...

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. ...

Three-Dimensionally Printed Microfluidic Cross-flow System for Ultrafiltration/Nanofiltration Membrane Performance Testing

Minimization and management of membrane fouling is a formidable challenge in diverse industrial processes and other practices that utilize membrane technology. Understanding the fouling process could lead to optimization and higher efficiency of membrane based filtration. Here we show the design and fabrication of an automated three-dimensionally (3-D) printed microfluidic cross-flow filtration system that can test up to 4 membranes in parallel. The microfluidic cells were printed using multi-material photopolymer 3-D printing technology, which used a transparent hard polymer for the microfluidic cell body and incorporated a thin rubber-like polymer layer, which prevents leakages during operation. The performance of ultrafiltration (UF), and nanofiltration (NF) membranes were tested and membrane fouling could be observed with a model foulant bovine serum albumin (BSA). Feed solutions containing BSA showed flux decline of the membrane. This protocol may be extended to measure fouling or biofouling with many other organic, inorganic or microbial containing solutions. The microfluidic design is especially advantageous for testing materials that are costly or only available in small quantities, for example polysaccharides, proteins, or lipids due to the small surface area of the membrane being tested. This modular system may also be easily expanded for high throughput testing of membranes.&#160;

Design and fabrication of a three-dimensionally (3-D) printed microfluidic cross-flow filtration system is demonstrated. The system is used to test performance and observe fouling of ultrafiltration and nanofiltration (thin film composite) membranes.

Minimization and management of membrane fouling is a formidable challenge in diverse industrial processes and other practices that utilize membrane ...

Microfluidic Platform with Multiplexed Electronic Detection for Spatial Tracking of Particles

Microfluidic processing of biological samples typically involves differential manipulations of suspended particles under various force fields in order to spatially fractionate the sample based on a biological property of interest. For the resultant spatial distribution to be used as the assay readout, microfluidic devices are often subjected to microscopic analysis requiring complex instrumentation with higher cost and reduced portability. To address this limitation, we have developed an integrated electronic sensing technology for multiplexed detection of particles at different locations on a microfluidic chip. Our technology, called Microfluidic CODES, combines Resistive Pulse Sensing with Code Division Multiple Access to compress 2D spatial information into a 1D electrical signal. In this paper, we present a practical demonstration of the Microfluidic CODES technology to detect and size cultured cancer cells distributed over multiple microfluidic channels. As validated by the high-speed microscopy, our technology can accurately analyze dense cell populations all electronically without the need for an external instrument. As such, the Microfluidic CODES can potentially enable low-cost integrated lab-on-a-chip devices that are well suited for the point-of-care testing of biological samples.

We demonstrate a microfluidic platform with an integrated surface electrode network that combines resistive pulse sensing (RPS) with code division multiple access (CDMA), to multiplex detection and sizing of particles in multiple microfluidic channels.

Microfluidic processing of biological samples typically involves differential manipulations of suspended particles under various force fields in order to ...

An Ultrasonic Tool for Nerve Conduction Block in Diabetic Rat Models

Nerve conduction block with a high intensity-focused ultrasound (HIFU) transducer has been performed in normal and diabetic animal models recently. HIFU can reversibly block the conduction of peripheral nerves without damaging the nerves while using an appropriate ultrasonic parameter. Temporary and partial block of the action potentials of nerves shows that HIFU has the potential to be a useful clinical treatment for pain relief. This work demonstrates the procedures for suppressing the action potentials of neuropathic nerves in diabetic rats in vivo using an HIFU transducer. The first step is to generate adult male diabetic neuropathic rats by streptozotocin (STZ) injection. The second step is to evaluate the peripheral diabetic neuropathy in STZ-induced diabetic rats by an electronic von Frey probe and a hot plate. The final step is to record in vivo extracellular action potentials of the nerve exposed to HIFU sonication. The method showed here may benefit the study of ultrasound analgesic applications.

This work presents the methodology of applying high intensity-focused ultrasound to block the action potentials of diabetic neuropathic nerves.

Nerve conduction block with a high intensity-focused ultrasound (HIFU) transducer has been performed in normal and diabetic animal models recently. HIFU ...

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

In the last decade, microfluidic techniques have been applied to study small animals, including the nematode Caenorhabditis elegans, and have proved useful as a convenient live imaging platform providing capabilities for precise control of experimental conditions in real time. In this article, we demonstrate live imaging of individual worms employing WormSpa, a previously-published custom microfluidic device. In the device, multiple worms are individually confined to separate chambers, allowing multiplexed longitudinal surveillance of various biological processes. To illustrate the capability, we performed proof-of-principle experiments in which worms were infected in the device with pathogenic bacteria, and the dynamics of expression of immune response genes and egg laying were monitored continuously in individual animals. The simple design and operation of this device make it suitable for users with no previous experience with microfluidic-based experiments. We propose that this approach will be useful for many researchers interested in longitudinal observations of biological processes under well-defined conditions.

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

In this article, we demonstrate live imaging of individual worms employing a custom microfluidic device. In the device, multiple worms are individually confined to separate chambers, allowing multiplexed longitudinal surveillance of various biological processes.

In the last decade, microfluidic techniques have been applied to study small animals, including the nematode Caenorhabditis elegans, and have proved ...

An Ultrahigh-throughput Microfluidic Platform for Single-cell Genome Sequencing

Sequencing technologies have undergone a paradigm shift from bulk to single-cell resolution in response to an evolving understanding of the role of cellular heterogeneity in biological systems. However, single-cell sequencing of large populations has been hampered by limitations in processing genomes for sequencing. In this paper, we describe a method for single-cell genome sequencing (SiC-seq) which uses droplet microfluidics to isolate, amplify, and barcode the genomes of single cells. Cell encapsulation in microgels allows the compartmentalized purification and tagmentation of DNA, while a microfluidic merger efficiently pairs each genome with a unique single-cell oligonucleotide barcode, allowing &gt;50,000 single cells to be sequenced per run. The sequencing data is demultiplexed by barcode, generating groups of reads originating from single cells. As a high-throughput and low-bias method of single-cell sequencing, SiC-seq will enable a broader range of genomic studies targeted at diverse cell populations.

Single-cell sequencing reveals genotypic heterogeneity in biological systems, but current technologies lack the throughput necessary for the deep profiling of community composition and function. Here, we describe a microfluidic workflow for sequencing &gt;50,000 single-cell genomes from diverse cell populations.

Sequencing technologies have undergone a paradigm shift from bulk to single-cell resolution in response to an evolving understanding of the role of ...