Authors thank Mr. Kotaro Okubo for the kind assistance with SEM imaging. This work was supported by the Japan Society for the Promotion of Science Grant-in-Aid for Basic Research (B) (20310069), Grant-in-Aid for Research Activity Start-up (25880021), and by research grants from the Kurata Memorial Hitachi Science and Technology Foundation and the Nippon Sheet Glass Foundation for Materials Science and Engineering.

包被的玻璃盖玻片1.准备二氧化钛<ol><li>号码使用金刚石划片盖玻片。这不仅有助于跟踪每个盖玻片的同时也确保了样品的正确的一面朝上。清洁的盖玻片，第一下运行DDH 2 O的，然后通过浸入它们食人鱼溶液（H 2 SO 4 :H 2 O 2 = 4:1）。 10分钟后，彻底冲洗盖玻片，8次双蒸2 O.在干燥N 2流盖玻片。 </li><li>在射频（RF）溅射系统设置的 TiO 2靶。附加的盖玻片上使用聚酰亚胺胶带将溅射设备的一个样品架。将样本保持在溅射室中。疏散室，直到压力达到2.0×10 -4帕。 </li><li>引入Ar气到腔室内，并设置在沉积压力为4.0毫乇。当保持闸板关闭时，逐渐增加RF功率至70瓦</li><li>打开快门和溅射15分钟，得到一个薄膜的厚度为120-150毫微米（ 图1:步骤A）。膜的生长速率需要被衍生为每个机。 </li></ol> 2.表面涂层与细胞排斥膜<ol><li>通过用O 2等离子体处理样品，以下由等离子体反应器的制造商提供的说明书亲水在TiO 2表面上。我们治疗的样品进行5分钟，在200瓦与O 2流量以100sccm。沉浸在样品中DDH 2 O的，并确认表面超亲水性。表面干燥彻底在N 2流。 </li><li>制备1mM的十八烷基三氯硅烷（OTS）溶液中加入39.6微升的OTS 100毫升甲苯。浸在1小时在室温下的溶液中的样品。进行此步骤内的N 2 -filled手套袋中（ 图1:步骤B）。 </li><li>要除去物理吸附的分子，sonicatE中的样品中的甲苯，丙酮，乙醇和DDH 2 O的，每次5分钟，在该顺序。冲洗样品四次新鲜的DDH 2 O的和干燥在N 2流的表面。表面应是疏水为100-110°的接触角。 </li></ol> 3. 前现场表面图案<ol><li>工作在层流罩，得出一些划痕，表面上的钻石划线。这些标记有助于跟踪处理过的区域，也使显微镜聚焦。通过浸渍在70％乙醇的样品5分钟消毒OTS涂覆的 TiO 2。然后用清水冲洗样品两次消毒的双蒸2 O. </li><li>放置在一个35毫米的培养皿的样品，并加2ml的PC12生长培养基（参见步骤4.2）。孵育超过3小时，在CO 2培养箱中（37℃）。这个程序的目的是让血清白蛋白吸收在表面上。吸附白蛋白抑制其他prote随后的吸附插件和细胞 （图1:步骤C）。 </li><li>在等待时，设置了倒置荧光显微镜。 <ol><li>打开弧光灯，插入UV滤光立方体，和物镜设置为20X。 </li><li>测量光强度 I（W厘米-2）用紫外线强度计，并计算辐照时间 t（秒）为一个在剂量d的（以J厘米-2）为:T = D / I。 例如，照射在为200J厘米-2的剂量使用的600毫瓦cm -2的光源，333秒的照射是必需的。 </li><li>使用一个阶段微米和关闭场光阑设置要被照射的区域的尺寸，例如，200微米。 </li></ol></li><li>在3小时温育后，补充用200μl的3.0毫克毫升-1 IV型胶原的培养基（COL-IV;最终浓度300微克毫升-1）。 </li><li>转移35毫米的菜显微镜阶段。发现划痕米柜，聚焦显微镜到样品表面上，并且照射UV光以为200J cm -2的（ 图1:步骤D）的剂量。紫外线照射的面积可以通过调整视场光阑的开口，或通过与仲裁几何形状的金属掩模代替场光阑被改变。 </li><li>替换为新鲜生长培养基的培养基（无COL-IV）中，并将样品放回到培养箱。 </li></ol> 4.细胞培养<ol><li> PC12细胞常规培养进行在胶原包被的塑料盘。 <ol><li>涂覆60mm的组织培养皿山口-Ⅳ，第一润湿表面与DDH 2 O的，吸所有双蒸2 O. </li><li>用双蒸2 O稀释原液（3毫克毫升-1）10X准备300微克毫升-1 COL-IV </li><li>加入200μl的300微克毫升-1每60毫米的菜COL-IV。让该溶液通过表面扩散。 </li><li>干吨他菜在层流罩约1小时。 </li><li>用4ml的Dulbecco磷酸盐缓冲盐水（D-PBS）的漂清表面的两倍。如果不立 ​​即使用的涂层菜，用4mL双蒸2 O.的涮菜一次吸气DDH 2 O的后，保存在孵化器的菜。尽量不要把它存放超过两个星期。 </li></ol></li><li> PC12细胞生长在该组成的生长培养基:Dulbecco改良的Eagle培养基（低葡萄糖）+ 10％胎牛血清+ 5％马血清+ 1％青霉素 - 链霉素溶液。孵育细胞在 CO 2培养箱中（37℃，5％的CO 2），并更换一半培养基每隔一天。传代细胞才达到汇合。 <ol><li>吸出生长培养基，并加入2毫升PBS +（D-PBS + 10毫克毫升-1牛血清白蛋白（BSA）+ 10mM EDTA的）预热至37℃。孵育5分钟，在37℃。轻轻敲击菜分离所有CELLS。 </li><li>收集细胞在15ml锥形管中。冲洗用3毫升新鲜的D-PBS，预热至37℃的菜。 </li><li>离心反应管，在150×g下4分钟。 </li><li>吸出上清液，并添加1毫升生长培养基。 </li><li>对于常规培养，分裂细胞1:3至1:5。 </li></ol></li><li>板块细胞对紫外线改性的OTS / TiO 2的样品，计数细胞密度，并在35毫米培养皿（ 图1:步骤E）添加3.0×10 5个细胞 。孵育在加湿培养箱的培养皿（37℃，5％CO 2）1-2天。既幼稚和神经生长因子（NGF）-differentiated PC12细胞可用于构图实验。对于NGF分化的7S-NGF 100纳克毫升-1电镀细胞到样品之前加入到生长培养基中数天。 NGF分化似乎增加了PC12细胞的粘附性和使处理更容易，尤其是在原位图案化Éxperiments。 </li></ol> 5. 原位表面图案<ol><li>经过1-2天的培养上易地修饰表面，确认细胞附着并只在紫外线照射区域增长。设置的显微镜，如步骤3.4中所述。 </li><li>将样品转移到含有生长培养基的新35毫米组织培养皿，100纳克毫升-1 NGF（在使用NGF分化的细胞的情况下），和100μg毫升-1 COL-IV。 </li><li>放在显微镜阶段的菜。找到相应的位置，并且照射UV光以为200J cm -2的（ 图1:步骤F，G）的 ​​剂量。新创建的许可区域表示与图1中以星号:步骤G </li><li>尽量在30分钟内完成一个样品的处理。照射后，将样品转移回未经COL-IV。 </li><li>携带与铜的菜时，要特别小心ltured细胞和从盘传送样品盘，以防止拆卸图案的细胞。 </li></ol><img alt="图1" src="/files/ftp_upload/53045/53045fig1highres.jpg" width="700" /> 图中，整个过程1示意图。详见说明书。 <a href="https://www.jove.com/files/ftp_upload/53045/53045fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

<ol>
	<li>Hughes, M. A., Brennan, P. M., Bunting, A. S., Shipston, M. J., Murray, A. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+Patterning+on+Photolithographically+Defined+Parylene-C:+SiO2+Substrates.">Cell Patterning on Photolithographically Defined Parylene-C: SiO2 Substrates.</a> J. Vis. Exp. (85), e50929 (2014).</li><li>Kleinfeld, D., Kahler, K. H., Hockberger, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+Outgrowth+of+Dissociated+Neurons+on+Patterned+Substrates.">Controlled Outgrowth of Dissociated Neurons on Patterned Substrates.</a> J. Neurosci. 8, 4098-4120 (1988).</li><li>Pensen, D., Heinz, W. F., Werbin, J. L., Hoh, J. H., Haviland, D. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electron+Beam+Patterning+of+Fibronectin+Nanodots+that+Support+Focal+Adhesion+Formation.">Electron Beam Patterning of Fibronectin Nanodots that Support Focal Adhesion Formation.</a> Soft Matter. 3, 1280-1284 (2007).</li><li>Tanii, T., 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=Application+of+Organosilane+Monolayer+Template+to+Quantitative+Evaluation+of+Cancer+Cell+Adhesive+Ability.">Application of Organosilane Monolayer Template to Quantitative Evaluation of Cancer Cell Adhesive Ability.</a> Jpn. J. Appl. Phys. 50, 06GL01 (2011).</li><li>Yamamoto, 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=In-Situ+Guidance+of+Individual+Neuronal+Processes+by+Wet+Femtosecond+Laser+Processing+of+Self-Assembled+Monolayers.">In-Situ Guidance of Individual Neuronal Processes by Wet Femtosecond Laser Processing of Self-Assembled Monolayers.</a> Appl. Phys. Lett. 99, 163701-1610 (2011).</li><li>Yamamoto, 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=Differential+Neurite+Outgrowth+is+Required+for+Axon+Specification+by+Cultured+Hippocampal+Neurons.">Differential Neurite Outgrowth is Required for Axon Specification by Cultured Hippocampal Neurons.</a> J. Neurochem. 123, 904-910 (2012).</li><li>Shen, K., Qi, J., Kam, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microcontact+Printing+of+Proteins+for+Cell+Biology.">Microcontact Printing of Proteins for Cell Biology.</a> J. Vis. Exp. (22), e1065 (2008).</li><li>Johnson, D. M., LaFranzo, N. A., Maurer, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Creating+Two-Dimensional+Patterned+Substrates+for+Protein+and+Cell+Confinement.">Creating Two-Dimensional Patterned Substrates for Protein and Cell Confinement.</a> J. Vis. Exp. (55), e3164 (2011).</li><li>Singhvi, R., 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=Engineering+Cell+Shape+and+Function.">Engineering Cell Shape and Function.</a> Science. 264, 696-698 (1126).</li><li>Chen, C. S., Mrksich, M., Huang, S., Whitesides, G. M., Ingber, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Geometric+Control+of+Cell+Life+and+Death.">Geometric Control of Cell Life and Death.</a> Science. 276, 1425-1428 (1997).</li><li>Degot, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+Visualization+and+Quantitative+Analysis+of+Drug+Effects+using+Micropatterned+Cells.">Improved Visualization and Quantitative Analysis of Drug Effects using Micropatterned Cells.</a> J. Vis. Exp. (46), e2514 (2010).</li><li>Nakanishi, 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=Photoactivation+of+a+Substrate+for+Cell+Adhesion+under+Standard+Fluorescence+Microscopes.">Photoactivation of a Substrate for Cell Adhesion under Standard Fluorescence Microscopes.</a> J. Am. Chem. Soc. 126, 16314-16315 (2004).</li><li>Kim, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Addressable+Micropatterning+of+Multiple+Proteins+and+Cells+by+Microscope+Projection+Photolithography+Based+on+a+Protein+Friendly+Photoresist.">Addressable Micropatterning of Multiple Proteins and Cells by Microscope Projection Photolithography Based on a Protein Friendly Photoresist.</a> Langmuir. 26, 12112-12118 (2010).</li><li>Deka, G., Okano, K., Kao, F. -. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+Photopatterning+of+Cells+In+Situ+by+Q-Switched+Neodymium-Doped+Yttrium+Ortho-Vanadate.">Dynamic Photopatterning of Cells In Situ by Q-Switched Neodymium-Doped Yttrium Ortho-Vanadate.</a> Laser. J. Biomed. Opt. 19, 011012 (2014).</li><li>Robertus, J., Browne, W. R., Feringa, B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+Control+over+Cell+Adhesive+Properties+using+Molecular-Based+Surface+Engineering+Strategies.">Dynamic Control over Cell Adhesive Properties using Molecular-Based Surface Engineering Strategies.</a> Chem. Soc. Rev. 39, 354-378 (2010).</li><li>Choi, I., Yeo, W. -. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-Assembled+Monolayers+with+Dynamicity+Stemming+from+(Bio)chemical+Conversions:+From+Construction+to+Application.">Self-Assembled Monolayers with Dynamicity Stemming from (Bio)chemical Conversions: From Construction to Application.</a> ChemPhysChem. 14, 55-69 (2013).</li><li>Nakanishi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Switchable+Substrates+for+Analyzing+and+Engineering+Cellular+Functions.">Switchable Substrates for Analyzing and Engineering Cellular Functions.</a> Chem. Asian J. 9, 406-417 (2014).</li><li>Yamamoto, 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=In+Situ+Modification+of+Cell-Culture+Scaffolds+by+Photocatalytic+Decomposition+of+Organosilane+Monolayers.">In Situ Modification of Cell-Culture Scaffolds by Photocatalytic Decomposition of Organosilane Monolayers.</a> Biofabrication. 6, 035021 (2014).</li><li>Sekine, K., Yamamoto, H., Kono, S., Ikeda, T., Kuroda, A., Tanii, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surface+Modification+of+Cell+Scaffold+in+Aqueous+Solution+using+TiO2+Photocatalysis+and+Linker+Protein+L2+for+Patterning+Primary+Neurons.">Surface Modification of Cell Scaffold in Aqueous Solution using TiO2 Photocatalysis and Linker Protein L2 for Patterning Primary Neurons.</a> e-J. Surf. Sci. Nanotech. 13, 213-218 (2015).</li><li>Arima, Y., Iwata, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+Surface+Functional+Groups+on+Protein+Adsorption+and+Subsequent+Cell+Adhesion+using+Self-Assembled+Monolayers.">Effects of Surface Functional Groups on Protein Adsorption and Subsequent Cell Adhesion using Self-Assembled Monolayers.</a> J. Mater. Chem. 17, 4079-4087 (2007).</li><li>Fujishima, A., Zhang, X., Tryk, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TiO2+Photocatalysis+and+Related+Surface+Phenomena.">TiO2 Photocatalysis and Related Surface Phenomena.</a> Surf. Sci. Rep. 63, 515-582 (2008).</li><li>Sigal, G. B., Mrksich, M., Whitesides, G. M. <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+Surface+Wettability+on+the+Adsorption+of+Proteins+and+Detergents.">Effect of Surface Wettability on the Adsorption of Proteins and Detergents.</a> J. Am. Chem. Soc. 120, 3464-3473 (1998).</li><li>Zhang, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Transparent+and+Photo-Patternable+Superhydrophobic+Film.">A Transparent and Photo-Patternable Superhydrophobic Film.</a> Chem. Commun. 2007, 4949-4951 (1039).</li><li>Kaech, S., Banker, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Culturing+Hippocampal+Neurons.">Culturing Hippocampal Neurons.</a> Nat. Protoc. 1, 2406-2415 (2006).</li></ol>

The authors have nothing to disclose.

在我们目前的协议， 二氧化钛膜通过射频磁控溅射形成。我们是有利于沉积的这种方法，因为它允许我们可重复使用亚纳米粗糙度调制成光催化二氧化钛薄膜。虽然溅射淀积过程是熟悉的材料科学家和电子工程师，它可能不是很访问生物学家。在这种情况下，旋涂的TiO 2膜 。将另外的选择23。在该方法中， 二氧化钛纳米颗粒溶解在溶剂中通过离心力铺展?...

半导体光刻工艺及其衍生物-诸如光刻1,2，电子束光刻3-6，和微接触印刷7-10 -现在已成为一个既定的工具在细胞生物学生长的活细胞在限定位置和几何形状。图案形成方法依赖于使用微制造基底的，由微岛细胞容许涂料在非允许的背景。例如基底用作模板图案的细胞。这些技术为我们提供了新的方法来设计的细胞和它们的功能在一个单和多细胞水平上，以提取细胞的固有特性，并增加细胞为基础的药物筛选11的吞吐量。 在细胞图案化的自由度度会大大增加，如果模板图案几何可以改变在原位 ， 即 ，当细胞是上一次一培养urface。常规方法的图案形成，不能直接应用于这里，因为它们处理样品中的气氛或真空。因此各 ​​种新型的表面改性技术已被提出，它是基于，例如，在光反应性化合物12,13或激光烧蚀5,14，仅举几例。所提出的方法已经由Robertus 等人已很好地审查 。15，以及最近由Choi 等人的 16和中西17。 在这里，在这篇文章中，我们描述了一种新的原位表面修饰，这需要对二氧化钛的优势有机分子的光催化分解（ 二氧化钛）表面18,19协议。在该方法中，TiO 2膜被插入在玻璃基板和接口单元中的有机薄膜之间，且有机薄膜通过局部地照射紫外线（UV）的分解原位光到感兴趣的区域（λ&lt;388纳米）。我们表明，新的协议可以用于创建细胞外基质蛋白和活细胞双方易地和原位微图案。 二氧化钛是生物相容的，化学稳定的，且光学透明的，特征，其中使得它友好引入在细胞培养实验。该协议提供了一个材料科学为基础的替代修改细胞培养支架的细胞培养环境。

<table><tbody><tr><td>Glass coverslip</td><td>Warner Instruments</td><td>CS-15R15</td><td>15 mm diameter, #1.5 thickness</td></tr><tr><td>Diamond scriber</td><td>Ogura Jewel Industry</td><td>D-Point Pen</td><td></td></tr><tr><td>RF sputtering system</td><td>ANELVA</td><td>SPC350</td><td></td></tr><tr><td>TiO&lt;sub&gt;2&lt;/sub&gt; sputtering target</td><td>Kojundo Chemical Lab</td><td>Titanium (IV) oxide, target</td><td>Purity, 99.9%</td></tr><tr><td>Plasma reactor</td><td>Yamato</td><td>PR301</td><td></td></tr><tr><td>n-octadecyltrichlorosilane (OTS)</td><td>Aldrich</td><td>104817</td><td></td></tr><tr><td>Toluene</td><td>Wako</td><td>204-01866</td><td></td></tr><tr><td>Tissue-culture dish (35&amp;#160;mm)</td><td>Greiner</td><td>627160</td><td></td></tr><tr><td>Tissue-culture dish (60&amp;#160;mm)</td><td>BD Falcon</td><td>353002</td><td></td></tr><tr><td>Type-IV collagen</td><td>Nitta Gelatin</td><td>Cellmatrix Type IV</td><td></td></tr><tr><td>D-PBS</td><td>Gibco</td><td>14190-144</td><td></td></tr><tr><td>Dulbecco&#039;s modified Eagle&#039;s medium (DMEM)</td><td>Gibco</td><td>11885-084</td><td></td></tr><tr><td>Fetal bovine serum</td><td>Gibco</td><td>12483-020</td><td>Heat-inactivate and pass through a 0.22&amp;#160;&amp;#956;m filter before use</td></tr><tr><td>Horse serum</td><td>Gibco</td><td>26050-088</td><td>Pass through a 0.22&amp;#160;&amp;#956;m filter before use</td></tr><tr><td>Penicillin-streptomycin (100x)</td><td>Nacalai tesque</td><td>26253-84</td><td></td></tr><tr><td>7S nerve growth factor (NGF)</td><td>Alomone Labs</td><td>N-130</td><td></td></tr><tr><td>Bovine serum albumin (BSA)</td><td>Sigma</td><td>A2153</td><td></td></tr><tr><td>EDTA</td><td>Dojindo</td><td>N001</td><td>Stock solution in 0.5 M</td></tr><tr><td>TiO&lt;sub&gt;2&lt;/sub&gt; nanoparticle</td><td>Tayca</td><td>TKD-701</td><td></td></tr></tbody></table>

photopatterning proteins and cells in aqueous environment using tio2 photocatalysis

吸附的二氧化钛（TiO 2的）的表面上的有机污染物可以通过光催化下的紫外线（UV）光分解。在这里，我们描述了一种新的协议采用二氧化钛光催化局部改变衬底表面的细胞亲和力。在这个实验中，一个薄的TiO 2膜为溅射镀膜在玻璃盖玻片，并在TiO 2表面随后改性以从十八烷基三氯硅烷（OTS）衍生的有机硅烷单层，其抑制细胞粘附。将样品浸在细胞培养基中，和聚焦UV光照射至一个八角形区域。当神经元细胞系PC12细胞铺在样本，细胞粘附只在紫外线照射的区域。进一步的研究表明这种表面改性也可以在原位进行，也就是说，即使当细胞生长在衬底上。适当的修改表面的所需的细胞外矩阵Protein胶原存在于在紫外线照射时的介质。这里提出的技术可以潜在地在图案形成多种细胞类型可以采用用于构建共培养系统或任意地操纵细胞培养下。

图2A示出了溅射沉积的 TiO 2膜的横截面扫描电子显微镜（SEM）图像。从观察，膜的厚度估计为大约150纳米。引人注目这里是所沉积的TiO 2膜的平整度。通过原子力显微镜（AFM）进一步分析显示，在表面的根均方（RMS）粗糙度为0.2纳米（ 图2B）。 当TiO 2的表面被改性以OTS，然后浸渍在含有血清的生长培养基中，血清白蛋白吸附到...

Watch this Scientific Journal Video about Photopatterning Proteins and Cells in Aqueous Environment Using TiO2 Photocatalysis at JoVE.com

Photopatterning Proteins and Cells in Aqueous Environment Using TiO2 Photocatalysis

We describe a protocol for modifying cell affinity of a scaffold surface in aqueous environment. The method takes advantage of titanium dioxide photocatalysis to decompose organic film in the photo-irradiated region. We show that it can be used to create microdomains of scaffolding proteins, both ex situ and in situ.

光图案蛋白质和细胞中的水环境中使用二氧化钛<sub&gt; 2</sub&gt;光催化

Organic contaminants adsorbed on the surface of titanium dioxide (TiO2) can be decomposed by photocatalysis under ultraviolet (UV) light. Here we describe ...

photopatterning-proteins-cells-aqueous-environment-using-tio2

Research

JoVE Journal

Bioengineering

Photopatterning Proteins and Cells in Aqueous Environment Using TiO2 Photocatalysis

7.8K 次观看.  Tohoku University. We describe a protocol for modifying cell affinity of a scaffold surface in aqueous environment. The method takes advantage of titanium dioxide photocatalysis to decompose organic film in the photo-irradiated region. We show that it can be used to create microdomains of scaffolding proteins, both ex situ and in situ.

视频: Photopatterning Proteins and Cells in Aqueous Environment Using TiO2 Photocatalysis

Preparation and Use of Photocatalytically Active Segmented Ag|ZnO and Coaxial TiO2-Ag Nanowires Made by Templated Electrodeposition

Photocatalytically active nanostructures require a large specific surface area with the presence of many catalytically active sites for the oxidation and reduction half reactions, and fast electron (hole) diffusion and charge separation. Nanowires present suitable architectures to meet these requirements. Axially segmented Ag|ZnO and radially segmented (coaxial) TiO2-Ag nanowires with a diameter of 200 nm and a length of 6-20 &#181;m were made by templated electrodeposition within the pores of polycarbonate track-etched (PCTE) or anodized aluminum oxide (AAO) membranes, respectively. In the photocatalytic experiments, the ZnO and TiO2 phases acted as photoanodes, and Ag as cathode. No external circuit is needed to connect both electrodes, which is a key advantage over conventional photo-electrochemical cells. For making segmented Ag|ZnO nanowires, the Ag salt electrolyte was replaced after formation of the Ag segment to form a ZnO segment attached to the Ag segment. For making coaxial TiO2-Ag nanowires, a TiO2 gel was first formed by the electrochemically induced sol-gel method. Drying and thermal annealing of the as-formed TiO2 gel resulted in the formation of crystalline TiO2 nanotubes. A subsequent Ag electrodeposition step inside the TiO2 nanotubes resulted in formation of coaxial TiO2-Ag nanowires. Due to the combination of an n-type semiconductor (ZnO or TiO2) and a metal (Ag) within the same nanowire, a Schottky barrier was created at the interface between the phases. To demonstrate the photocatalytic activity of these nanowires, the Ag|ZnO nanowires were used in a photocatalytic experiment in which H2 gas was detected upon UV illumination of the nanowires dispersed in a methanol/water mixture. After 17 min&#160;of illumination, approximately 0.2 vol% H2 gas was detected from a suspension of ~0.1 g of Ag|ZnO nanowires in a 50 ml 80 vol% aqueous methanol solution.

Preparation and Use of Photocatalytically Active Segmented Ag|ZnO and Coaxial TiO2-Ag Nanowires Made by Templated Electrodeposition

Procedures are outlined to prepare segmented and coaxial nanowires via templated electrodeposition in nanopores. As examples, segmented nanowires consisting of Ag and ZnO segments, and coaxial nanowires consisting of a TiO2 shell and a Ag core were made. The nanowires were used in photocatalytic hydrogen formation experiments.

制备和使用光催化活性分段银|氧化锌和二氧化钛同轴<sub&gt; 2</sub&gt;银纳米线制成由模板电

Photocatalytically active nanostructures require a large specific surface area with the presence of many catalytically active sites for the oxidation and ...

科研

工程学

Patterning Cells on Optically Transparent Indium Tin Oxide Electrodes

The ability to exercise precise spatial and temporal control over cell-surface interactions is an important prerequisite to the assembly of multi-cellular constructs serving as in vitro mimics of native tissues.  In this study, photolithography and wet etching techniques were used to fabricate individually addressable indium tin oxide (ITO) electrodes on glass substrates.  The glass substrates containing ITO microelectrodes were modified with poly(ethylene glycol) (PEG) silane to make them protein and cell resistive.  Presence of insulating PEG molecules on the electrode surface was verified by cyclic voltammetry employing potassium ferricyanide as a redox reporter molecule.  Importantly, the application of reductive potential caused desorption of the PEG layer, resulting in regeneration of the conductive electrode surface and appearance of typical ferricyanide redox peaks.    Application of reductive potential also corresponded to switching of ITO electrode properties from cell non-adhesive to cell-adhesive. Electrochemical stripping of PEG-silane layer from ITO microelectrodes allowed for cell adhesion to take place in a spatially defined fashion, with cellular patterns corresponding closely to electrode patterns.  Micropatterning of several cell types was demonstrated on these substrates.   In the future, the control of the biointerfacial properties afforded by this method will allow to engineer cellular microenvironments through the assembly of three or more cell types into a precise geometric configuration on an optically transparent substrate.

Non-fouling PEG silane monolayer was desorbed from individually addressable ITO electrodes on glass by application of a reductive potential. Electrochemical stripping of PEG-silane layer from ITO microelectrodes allowed for cell adhesion to take place in a spatially defined fashion, with cellular patterns corresponding closely to electrode patterns.

在光学透明的铟锡氧化物电极的图案细胞

The ability to exercise precise spatial and temporal control over cell-surface interactions is an important prerequisite to the assembly of multi-cellular ...

生物学

Ambient Method for the Production of an Ionically Gated Carbon Nanotube Common Cathode in Tandem Organic Solar Cells

A method of fabricating organic photovoltaic (OPV) tandems that requires no vacuum processing is presented. These devices are comprised of two solution-processed polymeric cells connected in parallel by a transparent carbon nanotubes (CNT) interlayer. This structure includes improvements in fabrication techniques for tandem OPV devices. First the need for ambient-processed cathodes is considered. The CNT anode in the tandem device is tuned via ionic gating to become a common cathode. Ionic gating employs electric double layer charging to lower the work function of the CNT electrode. Secondly, the difficulty of sequentially stacking tandem layers by solution-processing is addressed. The devices are fabricated via solution and dry-lamination in ambient conditions with parallel processing steps. The method of fabricating the individual polymeric cells, the steps needed to laminate them together with a common CNT cathode, and then provide some representative results are described. These results demonstrate ionic gating of the CNT electrode to create a common cathode and addition of current and efficiency as a result of the lamination procedure.

A method of fabricating, in ambient conditions, organic photovoltaic tandem devices in a parallel configuration is presented. These devices feature an air-processed, semi-transparent, carbon nanotube common cathode.

对于一个以离子门控碳纳米管共阴极的串联有机太阳能电池的生产环境法

A method of fabricating organic photovoltaic (OPV) tandems that requires no vacuum processing is presented. These devices are comprised of two ...

Photochemical Oxidative Growth of Iridium Oxide Nanoparticles on CdSe@CdS Nanorods

We demonstrate a procedure for the photochemical oxidative growth of iridium oxide catalysts on the surface of seeded cadmium selenide-cadmium sulfide (CdSe@CdS) nanorod photocatalysts. Seeded rods are grown using a colloidal hot-injection method and then moved to an aqueous medium by ligand exchange. CdSe@CdS nanorods, an iridium precursor and other salts are mixed and illuminated. The deposition process is initiated by absorption of photons by the semiconductor particle, which results with formation of charge carriers that are used to promote redox reactions. To insure photochemical oxidative growth we used an electron scavenger. The photogenerated holes oxidize the iridium precursor, apparently in a mediated oxidative pathway. This results in the growth of high quality crystalline iridium oxide particles, ranging from 0.5 nm to about 3 nm, along the surface of the rod. Iridium oxide grown on CdSe@CdS heterostructures was studied by a variety of characterization methods, in order to evaluate its characteristics and quality. We explored means for control over particle size, crystallinity, deposition location on the CdS rod, and composition. Illumination time and excitation wavelength were found to be key parameters for such control. The influence of different growth conditions and the characterization of these heterostructures are described alongside a detailed description of their synthesis. Of significance is the fact that the addition of iridium oxide afforded the rods astounding photochemical stability under prolonged illumination in pure water (alleviating the requirement for hole scavengers).

A protocol for the photochemical oxidative growth of small crystalline iridium oxide nanoparticles on the surface of CdSe@CdS seeded rod nanoparticles is presented.

铱氧化物纳米颗粒对硒化镉硫化镉@纳米棒光化学氧化增长

We demonstrate a procedure for the photochemical oxidative growth of iridium oxide catalysts on the surface of seeded cadmium selenide-cadmium sulfide ...

化学

Digital Printing of Titanium Dioxide for Dye Sensitized Solar Cells

Silicon solar cell manufacturing is an expensive and high energy consuming process. In contrast, dye sensitized solar cell production is less environmentally damaging with lower processing temperatures presenting a viable and low cost alternative to conventional production. This paper further enhances these environmental credentials by evaluating the digital printing and therefore additive production route for these cells. This is achieved here by investigating the formation and performance of a metal oxide photoelectrode using nanoparticle sized titanium dioxide. An ink-jettable material was formulated, characterized and printed with a piezoelectric inkjet head to produce a 2.6 &#181;m thick layer. The resultant printed layer was fabricated into a functioning cell with an active area of 0.25 cm2 and a power conversion efficiency of 3.5%. The binder-free formulation resulted in a reduced processing temperature of 250 &#176;C, compatible with flexible polyamide substrates which are stable up to temperatures of 350 &#730;C. The authors are continuing to develop this process route by investigating inkjet printing of other layers within dye sensitized solar cells.

This paper investigates the suitability of inkjet printing for the manufacturing of dye-sensitized solar cells. A binder-free TiO2 nanoparticle ink was formulated and printed onto a FTO glass substrate. The printed layer was fabricated into a cell with an active area of 0.25 cm2 and an efficiency of 3.5%.

钛白粉用于染料敏化太阳能电池的数码印刷

Silicon solar cell manufacturing is an expensive and high energy consuming process. In contrast, dye sensitized solar cell production is less ...

Cell Patterning on Photolithographically Defined 
Parylene-C: SiO2 Substrates

Cell patterning platforms support broad research goals, such as construction of predefined in vitro neuronal networks and the exploration of certain central aspects of cellular physiology. To easily combine cell patterning with Multi-Electrode Arrays (MEAs) and silicon-based &#8216;lab on a chip&#8217; technologies, a microfabrication-compatible protocol is required. We describe a method that utilizes deposition of the polymer parylene-C on SiO2&#160;wafers. Photolithography enables accurate and reliable patterning of parylene-C at micron-level resolution. Subsequent activation by immersion in fetal bovine serum (or another specific activation solution) results in a substrate in which cultured cells adhere to, or are repulsed by, parylene or SiO2 regions respectively. This technique has allowed patterning of a broad range of cell types (including primary murine hippocampal cells, HEK 293 cell line, human neuron-like teratocarcinoma cell line, primary murine cerebellar granule cells, and primary human glioma-derived stem-like cells). Interestingly, however, the platform is not universal; reflecting the importance of cell-specific adhesion molecules. This cell patterning process is cost effective, reliable, and importantly can be incorporated into standard microfabrication (chip manufacturing) protocols, paving the way for integration of microelectronic technology.

Cell Patterning on Photolithographically Defined 
Parylene-C: SiO2 Substrates

This protocol describes a microfabrication-compatible method for cell patterning on SiO2. A predefined parylene-C design is photolithographically printed on SiO2 wafers. Following incubation with serum (or other activation solution) cells adhere specifically to (and grow according to the conformity of) underlying parylene-C, whilst being repulsed by SiO2 regions.

在光刻技术定义的聚对二甲苯-C细胞图案：二氧化硅<sub&gt; 2</sub&gt;基板

Cell patterning platforms support broad research goals, such as construction of predefined in vitro neuronal networks and the exploration of certain ...

生物工程

Morphology Control for Fully Printable Organic&#8211;Inorganic Bulk-heterojunction Solar Cells Based on a Ti-alkoxide and Semiconducting Polymer

The photoactive layer of a typical organic thin-film bulk-heterojunction (BHJ) solar cell commonly uses fullerene derivatives as the electron-accepting material. However, fullerene derivatives are air-sensitive; therefore, air-stable material is needed as an alternative. In the present study, we propose and describe the properties of Ti-alkoxide as an alternative electron-accepting material to fullerene derivatives to create highly air-stable BHJ solar cells. It is well-known that controlling the morphology in the photoactive layer, which is constructed with fullerene derivatives as the electron acceptor, is important for obtaining a high overall efficiency through the solvent method. The conventional solvent method is useful for high-solubility materials, such as fullerene derivatives. However, for Ti-alkoxides, the conventional solvent method is insufficient, because they only dissolve in specific solvents. Here, we demonstrate a new approach to morphology control that uses the molecular bulkiness of Ti-alkoxides without the conventional solvent method. That is, this method is one approach to obtain highly efficient, air-stable, organic-inorganic bulk-heterojunction solar cells.

A method for fully printable, fullerene-free, highly air-stable, bulk-heterojunction solar cells based on Ti alkoxides as the electron acceptor and the electron-donating polymer fabrication is described here. Moreover, a method for controlling the morphology of the photoactive layer through the molecular bulkiness of the Ti-alkoxide units is reported.

基于一个钛醇盐和半导体聚合物为全可印刷有机 - 无机批量异质结太阳能电池形貌控制

The photoactive layer of a typical organic thin-film bulk-heterojunction (BHJ) solar cell commonly uses fullerene derivatives as the electron-accepting ...

Electrospinning of Photocatalytic Electrodes for Dye-sensitized Solar Cells

This work demonstrates a protocol to fabricate a fiber-based photoanode for dye-sensitized solar cells, consisting of a light-scattering layer made of electrospun titanium dioxide nanofibers (TiO2-NFs) on top of a blocking layer made of commercially available titanium dioxide nanoparticles (TiO2-NPs). This is achieved by first electrospinning a solution of titanium (IV) butoxide, polyvinylpyrrolidone (PVP), and glacial acetic acid in ethanol to obtain composite PVP/TiO2 nanofibers. These are then calcined at 500 &#176;C to remove the PVP and to obtain pure anatase-phase titania nanofibers. This material is characterized using scanning electron microscopy (SEM) and powder X-ray diffraction (XRD). The photoanode is prepared by first creating a blocking layer through the deposition of a TiO2-NPs/terpineol slurry on a fluorine-doped tin oxide (FTO) glass slide using doctor blading techniques. A subsequent thermal treatment is performed at 500 &#176;C. Then, the light-scattering layer is formed by depositing a TiO2-NFs/terpineol slurry on the same slide, using the same technique, and calcinating again at 500 &#176;C. The performance of the photoanode is tested by fabricating a dye-sensitized solar cell and measuring its efficiency through J-V curves under a range of incident light densities, from 0.25-1 Sun.

The overall goal of this project was to use electrospinning to fabricate a photoanode with improved performance for dye-sensitized solar cells.

染料敏化太阳能电池的光催化电极的电纺丝

This work demonstrates a protocol to fabricate a fiber-based photoanode for dye-sensitized solar cells, consisting of a light-scattering layer made of ...

The Effect of Interfacial Chemical Bonding in TiO2-SiO2 Composites on Their Photocatalytic NOx Abatement Performance

The chemical bonding of particulate photocatalysts to supporting material surfaces is of great importance in engineering more efficient and practical photocatalytic structures. However, the influence of such chemical bonding on the optical and surface properties of the photocatalyst and thus its photocatalytic activity/reaction selectivity behavior has not been systematically studied. In this investigation, TiO2 has been supported on the surface of SiO2 by means of two different methods: (i) by the in situ formation of TiO2 in the presence of sand quartz via a sol-gel method employing tetrabutyl orthotitanium (TBOT); and (ii) by binding the commercial TiO2 powder to quartz on a surface silica gel layer formed from the reaction of quartz with tetraethylorthosilicate (TEOS). For comparison, TiO2 nanoparticles were also deposited on the surfaces of a more reactive SiO2 prepared by a hydrolysis-controlled sol-gel technique as well as through a sol-gel route from TiO2 and SiO2 precursors. The combination of TiO2 and SiO2, through interfacial Ti-O-Si bonds, was confirmed by FTIR spectroscopy and the photocatalytic activities of the obtained composites were tested for photocatalytic degradation of NO according to the ISO standard method (ISO 22197&#8722;1). The electron microscope images of the obtained materials showed that variable photocatalyst coverage of the support surface can successfully be achieved but the photocatalytic activity towards NO removal was found to be affected by the preparation method and the nitrate selectivity is adversely affected by Ti-O-Si bonding.

The Effect of Interfacial Chemical Bonding in TiO2-SiO2 Composites on Their Photocatalytic NOx Abatement Performance

The focus of the present work is to establish means to generate and quantify levels of Ti-O-Si linkages and to correlate these with the photocatalytic properties of the supported TiO2.

界面化学键合在TiO中的作用<sub&gt; 2</sub&gt; -SiO<sub&gt; 2</sub&gt;复合材料对其光催化NOx消除性能的影响

The chemical bonding of particulate photocatalysts to supporting material surfaces is of great importance in engineering more efficient and practical ...

Developing Photosensitizer-Cobaloxime Hybrids for Solar-Driven H2 Production in Aqueous Aerobic Conditions

Developing photocatalytic H2 production devices is the one of the key steps for constructing a global H2-based renewable energy infrastructure. A number of photoactive assemblies have emerged where a photosensitizer and cobaloxime-based H2 production catalysts work in tandem to convert light energy into the H-H chemical bonds. However, the long-term instability of these assemblies and the need for hazardous proton sources have limited their usage. Here, in this work, we have integrated a stilbene-based organic dye into the periphery of a cobaloxime core via a distinct axial pyridine linkage. This strategy allowed us to develop a photosensitizer-catalyst hybrid structure with the same molecular framework. In this article, we have explained the detailed procedure of the synthesis of this hybrid molecule in addition to its comprehensive chemical characterization. The structural and optical studies have exhibited an intense electronic interaction between the cobaloxime core and the organic photosensitizer. The cobaloxime was active for H2 production even in the presence of water as the proton source. Here, we have developed a simple airtight system connected with an online H2 detector for the investigation of the photocatalytic activity by this hybrid complex. This photosensitizer-catalyst dyad present in the experimental setup continuously produced H2 once it was exposed in the natural sunlight. This photocatalytic H2 production by the hybrid complex was observed in aqueous/organic mixture media in the presence of a sacrificial electron donor under complete aerobic conditions. Thus, this photocatalysis measurement system along with the photosensitizer-catalyst dyad provide valuable insight for the development of next generation photocatalytic H2 production devices.

Developing Photosensitizer-Cobaloxime Hybrids for Solar-Driven H2 Production in Aqueous Aerobic Conditions

We have directly incorporated a stilbene-based organic dye into a cobaloxime core to generate a photosensitizer-catalyst dyad for photocatalytic H2 production. We have also developed a simple experimental setup for evaluating the light-driven H2 production by photocatalytic assemblies.

开发用于在水有条件条件下进行太阳能驱动 H2生产的光敏剂-钴氧混合剂

Developing photocatalytic H2 production devices is the one of the key steps for constructing a global H2-based renewable energy infrastructure. A number ...

光图案蛋白质和细胞中的水环境中使用二氧化钛

摘要

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此视频中的章节

制备和使用光催化活性分段银|氧化锌和二氧化钛同轴

在光学透明的铟锡氧化物电极的图案细胞

对于一个以离子门控碳纳米管共阴极的串联有机太阳能电池的生产环境法

铱氧化物纳米颗粒对硒化镉硫化镉@纳米棒光化学氧化增长

钛白粉用于染料敏化太阳能电池的数码印刷

在光刻技术定义的聚对二甲苯-C细胞图案：二氧化硅

基于一个钛醇盐和半导体聚合物为全可印刷有机 - 无机批量异质结太阳能电池形貌控制

染料敏化太阳能电池的光催化电极的电纺丝

界面化学键合在TiO中的作用

开发用于在水有条件条件下进行太阳能驱动 H₂生产的光敏剂-钴氧混合剂