This work was supported by the Shenzhen fundamental research funding (Grant No. JCYJ 20150630170146829, JCYJ20160531195439665 and JCYJ20160317152359560). The authors would like to thank Prof. Y. Chen at the Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences for supports.

1. Mold Fabrication
<ol>
	<li>Design two photomasks using a drawing software. Describe the outline of microchannel structure and use two separate layers for mask 1 and 2 in the same drawing file, so ensure all connections between different channels. Print different layers independently to chrome plate on the glass by a vendor with 1 &#181;m resolution. Ensure that the photomasks are dark with transparent designed structures, as a negative polarity. 
	Note: The mask 1 contains the dispersed phase i...

<ol>
	<li>Teh, S. Y., Lin, R., Hung, L. H., Lee, A. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Droplet+microfluidics.">Droplet microfluidics.</a> Lab on a chip. 8, 198-220 (2008).</li><li>Xiao, Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+multifunctional+NaYF4:Er3+,Yb3+/PEGDA+hybrid+microspheres:+NIR-light-activated+photopolymerization+and+drug+delivery.">Novel multifunctional NaYF4:Er3+,Yb3+/PEGDA hybrid microspheres: NIR-light-activated photopolymerization and drug delivery.</a> Chemical communications. 49, 1527-1529 (2013).</li><li>Pan, M., Kim, M., Blauch, L., Tang, S. K. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface-functionalizable+amphiphilic+nanoparticles+for+pickering+emulsions+with+designer+fluid-fluid+interfaces.">Surface-functionalizable amphiphilic nanoparticles for pickering emulsions with designer fluid-fluid interfaces.</a> RSC Adv. 6, 39926-39932 (2016).</li><li>Zhang, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+synthesis+of+rigid+nanovesicles+for+hydrophilic+reagents+delivery.">Microfluidic synthesis of rigid nanovesicles for hydrophilic reagents delivery.</a> Angewandte Chemie. 54, 3952-3956 (2015).</li><li>Shembekar, N., Chaipan, C., Utharala, R., Merten, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Droplet-based+microfluidics+in+drug+discovery,+transcriptomics+and+high-throughput+molecular+genetics.">Droplet-based microfluidics in drug discovery, transcriptomics and high-throughput molecular genetics.</a> Lab on a chip. 16, 1314-1331 (2016).</li><li>Soh, G. Y., Yeoh, G. H., Timchenko, V. <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+on+the+velocity+fields+during+droplet+formation+in+a+microfluidic+T-junction.">Numerical investigation on the velocity fields during droplet formation in a microfluidic T-junction.</a> Chemical Engineering Science. 139, 99-108 (2016).</li><li>Chiarello, E., Derzsi, L., Pierno, M., Mistura, G., Piccin, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+Oil+Droplets+in+a+Non-Newtonian+Liquid+Using+a+Microfluidic+T-Junction.">Generation of Oil Droplets in a Non-Newtonian Liquid Using a Microfluidic T-Junction.</a> Micromachines. 6, 1825-1835 (2015).</li><li>Moyle, T. M., Walker, L. M., Anna, S. 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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Water-in-Water+Droplets+by+Passive+Microfluidic+Flow+Focusing.">Water-in-Water Droplets by Passive Microfluidic Flow Focusing.</a> Analytical chemistry. 88, 3982-3989 (2016).</li><li>Zhu, P., Tang, X., Wang, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Droplet+generation+in+co-flow+microfluidic+channels+with+vibration.">Droplet generation in co-flow microfluidic channels with vibration.</a> Microfluidics and Nanofluidics. , 20 (2016).</li><li>Kim, S. H., Kim, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+formation+of+double-emulsion+drops+in+sudden+expansion+channels.">Controlled formation of double-emulsion drops in sudden expansion channels.</a> Journal of colloid and interface science. 415, 26-31 (2014).</li><li>Jeong, W. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+generation+of+submicron+emulsion+droplets+via+highly+stable+tip-streaming+mode+in+microfluidic+devices.">Controlled generation of submicron emulsion droplets via highly stable tip-streaming mode in microfluidic devices.</a> Lab on a chip. 12, 1446-1453 (2012).</li><li>Peng, L., Yang, M., Guo, S. S., Liu, W., Zhao, X. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+interfacial+tension+on+droplet+formation+in+flow-focusing+microfluidic+device.">The effect of interfacial tension on droplet formation in flow-focusing microfluidic device.</a> Biomedical microdevices. 13, 559-564 (2011).</li><li>Nunes, J. K., Tsai, S. S., Wan, J., Stone, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dripping+and+jetting+in+microfluidic+multiphase+flows+applied+to+particle+and+fiber+synthesis.">Dripping and jetting in microfluidic multiphase flows applied to particle and fiber synthesis.</a> J Phys D Appl Phys. , 46 (2013).</li><li>Anna, S. L., Mayer, H. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microscale+tipstreaming+in+a+microfluidic+flow+focusing+device.">Microscale tipstreaming in a microfluidic flow focusing device.</a> Physics of Fluids. 18, 121512 (2006).</li><li>Liu, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+synthesis+of+QD-encoded+PEGDA+microspheres+for+suspension+assay.">Microfluidic synthesis of QD-encoded PEGDA microspheres for suspension assay.</a> J. Mater. Chem. B. 4, 482-488 (2016).</li><li>Richardson, W. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bayesian-based+iterative+method+of+image+restoration.">Bayesian-based iterative method of image restoration.</a> Journal of the Optical Society of America. 62, 55-59 (1972).</li><li>Mukhopadhyay, P., Chaudhuri, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+survey+of+Hough+Transform.">A survey of Hough Transform.</a> Pattern Recognition. 48, 993-1010 (2015).</li><li>Ward, T., Faivre, M., Stone, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drop+production+and+tip-streaming+phenomenon+in+a+microfluidic+flow-focusing+device+via+an+interfacial+chemical+reaction.">Drop production and tip-streaming phenomenon in a microfluidic flow-focusing device via an interfacial chemical reaction.</a> Langmuir: the ACS journal of surfaces and colloids. 26, 9233-9239 (2010).</li><li>Anna, S. L., Bontoux, N., Stone, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Formation+of+dispersions+using+"flow+focusing"+in+microchannels.">Formation of dispersions using "flow focusing" in microchannels.</a> Applied Physics Letters. 82, 364 (2003).</li><li>Kim, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+production+of+emulsion+drops+using+an+electric+field+in+a+flow-focusing+microfluidic+device.">Controlled production of emulsion drops using an electric field in a flow-focusing microfluidic device.</a> Applied Physics Letters. 91, 133106 (2007).</li><li>Dangla, R., Gallaire, F., Baroud, C. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microchannel+deformations+due+to+solvent-induced+PDMS+swelling.">Microchannel deformations due to solvent-induced PDMS swelling.</a> Lab on a chip. 10, 2972-2978 (2010).</li><li>Chiu, Y. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Universally+applicable+three-dimensional+hydrodynamic+microfluidic+flow+focusing.">Universally applicable three-dimensional hydrodynamic microfluidic flow focusing.</a> Lab on a chip. 13, 1803-1809 (2013).</li><li>Oh, K. W., Lee, K., Ahn, B., Furlani, E. 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The generation of droplets in the flow-focusing mode using 2D and semi-3D microfluidic device has previously been developed in a variety of reports8,9,15,19,20,21. In these systems, the aqueous liquid that could not be solidified was chosen as the dispersed phase, such as deionized water8,<s...

Droplet-based microfluidic systems have the ability to produce highly monodisperse droplets from nanometer to micrometer diameter range1 and hold great potential in the high-throughput drug discovery2, synthesis of biomolecules3,4, and the diagnostic testing5. Due to the unique advantages of smaller droplets, such as the greater surface area to volume ratio and the large-scale applications with consuming a few microliters of sample, the technology has attracted extensive interest in a broad range of fields. The emulsification of two immiscible liquids is one of the most typical methods to generate droplet. In previous reports in the field, researchers have developed a variety of different droplet formation geometries, including T-junction, flow-focusing and co-flowing geometries. In the T-junction geometry, the dispersed phase is delivered through a perpendicular channel into the main channel, in which the continuous phase flows6,7. In the typical two-dimensional (2D) flow-focusing8,9&#160;geometry, the dispersed phase flow is sheared from the lateral; and for the co-flowing geometry10,11, on the other hand, a capillary introducing the dispersed phase flow is placed co-axially inside a bigger capillary for co-flowing geometry, so that the dispersed phase flow is sheared from all directions.
The droplet size is controlled by adjusting channel size and flow rate ratio, and the minimum size produced by co-flowing or T-junction is limited to dozens of micrometers. For flow-focusing droplet formation system, three modes of droplet breakup form by adjusting the pressure ratio of two-phase and surfactant concentration, including the dripping regime, the jetting regime, and tip-streaming15. Tip-streaming mode is also called thread formation, and the appearance of a thin the thread drawing out from the tip of dispersed phase flow cone will be observed.Previous studies have demonstrated droplets less than a few micrometers could be generated though tip-streaming process in 2D or semi-3D flow-focusing device8,12. However, as an aqueous solution containing a very low concentration of PEGDA was used as the dispersed phase, the shrinkage ratio of PEGDA particles was about 60% of the original droplets in diameter after photo-polymerization, while PEGDA without dilution as the dispersed phase led to unstable tip-streaming mode12. Interfacial tension is an important parameter of emulsion process and it will decrease due to the addition of the surfactant into the continuous phase liquid,leading to decrease in droplet size, higher generation frequency13, highly curved tip, and preventing instability14. Furthermore, when the bulk surfactant concentration is much higher than the critical micelle concentration, the interfacial tension is approximately invariable in the saturated state13 and the tip-streaming mode can occur15.
Based on the above observations, in this paper, we developed a facile approach for PEGDA droplets generation using a semi-3D flow-focusing microfluidic device, fabricated by multi-layer soft lithography method. Different from the typical 2D flow-focusing device, the semi-3D flow-focusing device has a shallow dispersed phase channel and a deep continuous phase channel, so that the dispersed phase can be sheared from up and down beside lateral. This provides larger adjusting range for flow-focusing mode by reducing the energy and pressure required for droplet breakup. Different from the previous report12, the dispersed phase is pure PEGDAcontaining photo-initiator, making sure that the shrinkage ratio of PEGDA particles is lower than 10%16; and the continuous phase is the mixture of hexadecane dissolving with a high bulk concentration of the silicone-based nonionic surfactant. Size-controllable and uniform droplets were produced by adjusting the pressure ratio of two phases. The diameter of the droplets changes from 80 &#181;m to 1 &#181;m as the droplet breakup processes changes from the jetting mode to tip-streaming mode. In addition, the PEGDA particle was synthesized through photo-polymerization process under UV exposure. The droplet generation microfluidic system with ease of fabrication will provide more possibilities for biological applications.

<table border="0" cellpadding="0" cellspacing="0" style="width:572px;" width="573">
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	</colgroup>
	<tbody>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">Silicon wafer</td>
			<td style="width:71px;">Huashi Co., Ltd</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">SU-8 2025, 2100</td>
			<td style="width:71px;">Microchem Co.</td>
			<td style="width:119px;">Y111069</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">SU-8 developer</td>
			<td style="width:71px;">Microchem Co.</td>
			<td style="width:119px;">Y020100</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="91">
			<td height="91" style="height:91px;width:216px;">Chromium mask</td>
			<td style="width:71px;">Qingyi Precision Mask Making Co., Ltd</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">polydimethylsiloxane(PDMS)</td>
			<td style="width:71px;">Dow Corning</td>
			<td style="width:119px;">Sylgard 184</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">poly(ethylene glycol) diacrylate (PEGDA)</td>
			<td style="width:71px;">Sigma</td>
			<td style="width:119px;">26570-48-9</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">2-hydroxy-40-(2-hydroxyethoxy)-2-methylpropiophenone</td>
			<td style="width:71px;">TCI</td>
			<td style="width:119px;">H1361-5G</td>
			<td style="width:167px;">photoinitiator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Hexadecane</td>
			<td style="width:71px;">Sigma</td>
			<td style="width:119px;">544-76- 3</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">ABIL EM 90</td>
			<td style="width:71px;">CHT</td>
			<td style="width:119px;">144243-53-8</td>
			<td style="width:167px;">surfactants</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Rhodamine B</td>
			<td style="width:71px;">Aladdin</td>
			<td style="width:119px;">81-88-9</td>
			<td style="width:167px;">fluorescent dye</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Spin Coater</td>
			<td style="width:71px;"></td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Lithography machine</td>
			<td style="width:71px;"></td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Automatic ointment agitator</td>
			<td style="width:71px;">Thinky</td>
			<td style="width:119px;">ARV-310</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Oven</td>
			<td style="width:71px;">BluePard</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Optical microscope</td>
			<td style="width:71px;">OLYMPUS</td>
			<td style="width:119px;">IX71</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">High-speed camera</td>
			<td style="width:71px;">Hamamatsu, Japan</td>
			<td style="width:119px;">ORCA-flash</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:216px;">MAESFLO Microfluidic Fluid Control System</td>
			<td style="width:71px;">FLUIGENT</td>
			<td style="width:119px;">MFCS-EZ</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:216px;">UV lamp</td>
			<td style="width:71px;">FUTANSI</td>
			<td style="width:119px;"></td>
			<td style="width:167px;">365 nm UV light, 8000 MW/CM2</td>
		</tr>
	</tbody>
</table>

generation of size-controlled poly (ethylene glycol) diacrylate droplets via semi-3-dimensional flow focusing microfluidic devices

Uniform and size-controllable poly (ethylene glycol) diacrylate (PEGDA) droplets could be produced via the flow focusing process in a microfluidic device. This paper proposes a semi-three-dimensional (semi-3D) flow-focusing microfluidic chip for droplet formation. The polydimethylsiloxane (PDMS) chip was fabricated using the multi-layer soft lithography method. Hexadecane containing surfactant was used as the continuous phase, and PEGDA with the ultraviolet (UV) photo-initiator was the dispersed phase. Surfactants allowed the local surface tension to drop and formed a more cusped tip which promoted breaking into tiny micro-droplets. As the pressure of dispersed phase was constant, the size of droplets became smaller with increasing continuous phase pressure before dispersed phase flow was broken off. As a result, droplets with size variation from 1 &#181;m to 80 &#181;m in diameter could be selectively achieved by changing the pressure ratio in two inlet channels, and the average coefficient of variation was estimated to be below 7%. Furthermore, droplets could turn into micro-beads by UV exposure for photo-polymerization. Conjugating biomolecules on such micro-beads surface have many potential applications in the fields of biology and chemistry.

The semi-3D flow-focusing microfluidic chip was fabricated using multi-layer soft lithography techniques as described above. The fabrication process and results for master mold in the protocolare shown in Figure 2. The first layer, which provides a 65 &#181;m wide channel for introducing the dispersed phase and a 50 &#181;m wide orifice (Figure 2a), is 20 &#181;m in thickness. An addition 130 &#181;m thickness la...

Watch this Scientific Journal Video about Generation of Size-controlled Poly ethylene Glycol Diacrylate Droplets via Semi-3-Dimensional Flow Focusing Microfluidic Devices at JoVE.com

Generation of Size-controlled Poly ethylene Glycol Diacrylate Droplets via Semi-3-Dimensional Flow Focusing Microfluidic Devices

Here, we present a protocol to illustrate the fabrication processes and verifying experiments of a semi-three-dimensional (semi-3D) flow-focusing microfluidic chip for droplet formation.

Generation of Size-controlled Poly (ethylene Glycol) Diacrylate Droplets via Semi-3-Dimensional Flow Focusing Microfluidic Devices

Uniform and size-controllable poly (ethylene glycol) diacrylate (PEGDA) droplets could be produced via the flow focusing process in a microfluidic device. ...

This method can help to answer the key questions in the microfluidic droplet formation field, including fabricating the semi-three dimensional flow focusing PDMS chip, and producing the pure PGD droplets. The main advantage of this tactic is that the semi-3D flow focusing device provides larger testing grounds for flow focusing mold. Pure PGD makes sure that the ratio of PGD particles is lower than 10%The implications of this technique can be extended to many biochemical implications due to the greater surface area to volume ratio.Nozzle, large-scale applications will be benefit for this technique with consuming a few microliters of sample. Now this method can provide insight into PGD droplet formation. It can also be applied to other systems such as gas-equipped flow focusing analyzing.Generally, individuals new to this method will struggle because of clogging in the orifice and the instability of tip streaming mold. Of this method is critical, as semi-3d flow focusing chip fabrication is difficult to learn because the alignment of top and bottom PDMS layers to fabricate semifluidic chip by hands is not reviewed. Design the two photo masks using standard techniques.Use two separate layers for mask one and two in the same drawing file to ensure that all connections line up between the different channels. Mask one contains the dispersed phased inlet channel and an orifice, while mask two contains the continuous phase inlet channel, the filter, and the outlet. Use the two alignment marks and print the different layers independently to a chrome plate on glass.Next, in a designated photolithography lab, place a clean, 3-inch diameter silicon wafer on a spin coater and turn on the vacuum to affix the wafer to the spin chuck. Spin coat two to three milliliters of SU-8 2025 negative photoresist onto the wafer for 10 seconds at 1, 000 rpm. Then change the speed to 3, 000 rpms for 30 seconds.This will provide a layer thickness of 20 microns for the first layer. Soft bake the first layer by placing it on a 95 degrees celsius hot plate for six minutes. And then, remove the wafer and let it cool back to room temperature.Expose the soft-baked wafer to mask number one under a collimated UV light for 18 seconds. Following UV exposure, post bake the wafer on a 95 degrees celsius hot plate for six minutes. Afterwards, allow the wafer to cool back to room temperature.Repeat the spincoating process a second time using two to three milliliters of SU-8 2100 negative photoresist. Spin the wafer for 10 seconds at 1, 000 rpm, and then at 2, 000 rpms for 30 seconds. This provides a second layer thickness of 130 microns.After soft baking the wafer on a 95 degrees celsius hot plate for 35 minutes, line up mask number two on the second layer photoresist, and expose it to UV light for 30 seconds. Then, post bake it on a 95 degree celsius hot plate for seven minutes. Once cooled back to room temperature, develop the wafer by immersing in a stirred bath of 50 milliliters of propylene glycol methyl ether acetate until features become clear on the wafer.Then, wash the wafer with ethyl alcohol. Finally, place the wafer on the thermostatic platform and hard bake it at 95 degrees celsius for two hours. Mix a PDMS monomer and its curing agent for four minutes at a 10:1 ratio for the top layer and an 8:1 ratio for the bottom layer.Using an automatic ointment agitator, place the silicon wafer into a 90 millimeter Petri dish, and then pour the 8:1 ratio PDMS mixture over the mold to a thickness of two to three millimeters. Then, degas the PDMS in a vacuum chamber until all the bubbles disappear. Place the Petri dish in an oven and cure the PDMS at 80 degrees celsius for one hour.Once cooled back to room temperature, use a scalpel to cut the device at least three millimeters away from the features, and slowly peel the PDMS layer from the silicon wafer. Then, punch the dispersed phase inlet, the continuous phase inlet, and the outlet in the top PDMS layer using a 0.75 millimeter diameter punch. Clean the PDMS surface with adhesive tape to remove any dust particles.Then, place both the top and bottom PDMS layers simultaneously into a plasma cleaner, and expose to plasma for two minutes at 300 watts. Immediately following plasma exposure, place the top layer on the bottom layer and slide the surfaces relatively until features are aligned viewing through a stereo microscope. Cure the device in an oven at 120 degrees celsius for one day to enhance its strength and complete the bonding.To begin, prepare the continuous phase solution by dissolving 18%of a silicone-based non-ionic surfactant in hexadecane. Next, prepare the solution of the dispersed phase by starting with a hydrophilic peg diacrylate, one millimole per liter of a fluorescent dye, and adding five milligrams per milliliter of a photo initiator. Fill the one milliliter reservoirs of a pneumatic pressure controller with the continuous phase.Then, fill a 200 microliter gel-loading tip with the dispersed phase. Place the semi-3D microfluidic device on the stage of an inverted optical microscope equipped with a high speed camera. Next, connect a fluorinated ethylene propylene tube to the punched hole of the continuous phase by first attaching it to a short, stainless steel tube, inserting the tube into the end of a gel loading tip, and placing the tip into the punched hole of the dispersed phase.Then, insert a 20 cm long FEP tube into the outlet of the device, and place the end into a Petri dish. Focus the inverted microscope on a region that contains the two phase intersection, the orifice region, and the downstream channel. Then, set pressures of the two phases using a connected pneumatic pressure controller.Set the dispersed phase at 15 millibar, and the continuous phase at 30 millibar. Wait three minutes for the pressure to stabilize, and stable fluid flow is reached, carrying no bubbles or PDMS residue. Next, set the pressure of the dispersed phase as the base level pressure of the system to 45 millibar.Increase the pressure of the continuous phase until the emulsion breakup mode is changed from jetting to the tip streaming mode. Then, wait five minutes for it to stabilize. Once set up, place the tube end connecting the outlet hole to Petri dish, and collect the droplets.Expose them to UV light where they will rapidly solidify into spheres. Because of the difference of depth of the dispersed phase channel, and the continuous phase channel, the dispersed phase flow is squeezed on all directions by the continuous phase flow. As a result, symmetric conical liquid tip forms to produce droplets continuously.The size of droplets is changed by the pressure ratio of dispersed and continuous phases flow. Here, the pressure of the continuous phase is modified to affect the sheering force so that the droplet breakup processes change from the jetting mode to the tip streaming mode. The droplets are solidified by photopolymerization forming particles upon exiting the chip.The droplets shown here were formed with different pressure ratios. The resulting droplet radius has an inverse relationship with the continuous phase to dispersed phase pressure ratio. This technique paves the way for researchers in the field of biological applications and provides a powerful tool for drug discovery, synthesis of biomolecules and the diagnostic testing.After watching this video, you should have a good understanding of how to fabricate semi-three dimensional microfluidic chips, and how to produce a few micro PGD droplets. Don't forget that working with photoresist PGMEA has a can be extremely hazardous and precautions such as wearing disposable gloves and a mask should always be taken while performing this procedure.

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7.6K 조회수.  Tsinghua University. This method can help to answer the key questions in the microfluidic droplet formation field, including fabricating the semi-three dimensional flow focusing PDMS chip, and producing the pure PGD droplets. The main advantage of this tactic is that the semi-3D flow focusing device provides larger testing grounds for flow focusing mold. Pure PGD makes sure that the ratio of PGD particles is lower than 10%The implications of this technique can be extended to many biochemical implications due to t...