Name: generation of scalable, metallic high-aspect ratio nanocomposites in a biological liquid medium
Uploaded: 2015-07-08T15:00:00
Description: Watch this Scientific Journal Video about Generation of Scalable, Metallic High-Aspect Ratio Nanocomposites in a Biological Liquid Medium at JoVE.com

The authors would like to acknowledge the technical assistance of Alfred Gunasekaran in electron microscopy studies at the Institute of Micromanufacturing at Louisiana Tech University, and Dr. Jim McNamara for assistance with additional microscopy studies. The work described was supported in part by Louisiana board of Regents PKSFI Contract No. LEQSF (2007-12)-ENH-PKSFI-PRS-04 and the James E. Wyche III Endowed Professorship from Louisiana Tech University (to M.D.).

1.规划实验<ol><li>确定所需的合成铜纳米复合材料的体积。在此基础上，选择一些体积小烧瓶（25 平方厘米），或在准备材料如下所示大瓶。 </li><li>对于此合成中，使用一个37℃培养箱，用5％的CO 2和至少40％的湿度。确保这种培养箱可用，并且，它不会被重复干扰比合成的周期（大约24小时）。 注意：反复开闭的保育箱的肯定会引起温度的波动，这可能会导致该纳米复合材?...

<ol>
	<li>Klinman, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+copper-enzyme+family+of+dopamine+beta-monooxygenase+and+peptidylglycine+alpha-hydroxylating+monooxygenase:+resolving+the+chemical+pathway+for+substrate+hydroxylation.">The copper-enzyme family of dopamine beta-monooxygenase and peptidylglycine alpha-hydroxylating monooxygenase: resolving the chemical pathway for substrate hydroxylation.</a> The Journal of biological chemistry. 281, 3013-3016 (2006).</li><li>Uauy, R., Olivares, M., Gonzalez, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Essentiality+of+copper+in+humans.">Essentiality of copper in humans.</a> The American journal of clinical nutrition. 67, 952S-959S (1998).</li><li>Karlsson, H. L., Cronholm, P., Gustafsson, J., Copper Moller, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=oxide+nanoparticles+are+highly+toxic:+a+comparison+between+metal+oxide+nanoparticles+and+carbon+nanotubes.">oxide nanoparticles are highly toxic: a comparison between metal oxide nanoparticles and carbon nanotubes.</a> Chemical research in toxicology. 21, 1726-1732 (2008).</li><li>Parekh, G., 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=Layer-by-layer+nanoencapsulation+of+camptothecin+with+improved+activity.">Layer-by-layer nanoencapsulation of camptothecin with improved activity.</a> International journal of pharmaceutics. 465, 218-227 (2014).</li><li>Harrington, M. J., Masic, A., Holten-Andersen, N., Waite, J. H., Fratzl, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Iron-clad+fibers:+a+metal-based+biological+strategy+for+hard+flexible+coatings.">Iron-clad fibers: a metal-based biological strategy for hard flexible coatings.</a> Science. 328, 216-220 (2010).</li><li>Keyson, D., 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=CuO+urchin-nanostructures+synthesized+from+a+domestic+hydrothermal+microwave+method.">CuO urchin-nanostructures synthesized from a domestic hydrothermal microwave method.</a> Materials Research Bulletin. 43, 771-775 (2008).</li><li>Liu, B., Zeng, 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=Mesoscale+organization+of+CuO+nanoribbons:+formation+of+'dandelions'.">Mesoscale organization of CuO nanoribbons: formation of 'dandelions'.</a> J Am Chem Soc. 126, 8124-8125 (2004).</li><li>Peng, 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=Controllable+synthesis+of+self-assembled+Cu2S+nanostructures+through+a+template-free+polyol+process+for+the+degradation+of+organic+pollutant+under+visible+light.">Controllable synthesis of self-assembled Cu2S nanostructures through a template-free polyol process for the degradation of organic pollutant under visible light.</a> Materials Research Bulletin. 44, 1834-1841 (2009).</li><li>Deodhar, S., Huckaby, J., Delahoussaye, M., DeCoster, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Aspect+Ratio+Bio-Metallic+Nanocomposites+for+Cellular+Interactions.">High-Aspect Ratio Bio-Metallic Nanocomposites for Cellular Interactions.</a> IOP Conference Series: Materials Science and Engineering. 64, 012014 (2014).</li><li>Montes-Burgos, I., 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=Characterisation+of+nanoparticle+size+and+state+prior+to+nanotoxicological+studies.">Characterisation of nanoparticle size and state prior to nanotoxicological studies.</a> Journal of Nanoparticle Research. 12, 47-53 (2010).</li><li>Wiogo, H. T., Lim, M., Bulmus, V., Yun, J., Amal, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stabilization+of+magnetic+iron+oxide+nanoparticles+in+biological+media+by+fetal+bovine+serum+(FBS).">Stabilization of magnetic iron oxide nanoparticles in biological media by fetal bovine serum (FBS).</a> Langmuir. 27, 843-850 (2011).</li><li>Yunker, P. J., Still, T., Lohr, M. A., Yodh, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Suppression+of+the+coffee-ring+effect+by+shape-dependent+capillary+interactions.">Suppression of the coffee-ring effect by shape-dependent capillary interactions.</a> Nature. 476, 308-311 (2011).</li><li>Kahler, H., Lloyd Jr, B., Eden, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electron+Microscopic+and+Other+Studies+on+a+Copper&#8211;Cystine+Complex.">Electron Microscopic and Other Studies on a Copper&#8211;Cystine Complex.</a> The Journal of Physical Chemistry. 56, 768-770 (1952).</li><li>Furia, E., Sindona, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Complexation+of+L-cystine+with+metal+cations.">Complexation of L-cystine with metal cations.</a> Journal of Chemical &amp; Engineering Data. 55, 2985-2989 (2010).</li><li>Hawkins, C., Perrin, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polynuclear+Complex+Formation.+II.+Copper+(II)+with+Cystine+and+Related+Ligands.">Polynuclear Complex Formation. II. Copper (II) with Cystine and Related Ligands.</a> Inorganic Chemistry. 2, 843-849 (1963).</li><li>Hallman, P., Perrin, D., Watt, A. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+computed+distribution+of+copper+(II)+and+zinc+(II)+ions+among+seventeen+amino+acids+present+in+human+blood+plasma.">The computed distribution of copper (II) and zinc (II) ions among seventeen amino acids present in human blood plasma.</a> Biochem. J. 121, 549-555 (1971).</li><li>Jensen, L. S., Maurice, D. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+sulfur+amino+acids+on+copper+toxicity+in+chicks.">Influence of sulfur amino acids on copper toxicity in chicks.</a> The Journal of nutrition. 109, 91-97 (1979).</li><li>Lee, Y., Choi, J. R., Lee, K. J., Stott, N. E., Kim, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large-scale+synthesis+of+copper+nanoparticles+by+chemically+controlled+reduction+for+applications+of+inkjet-printed+electronics.">Large-scale synthesis of copper nanoparticles by chemically controlled reduction for applications of inkjet-printed electronics.</a> Nanotechnology. 19, 415604 (2008).</li><li>Hume, 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=Engineered+coiled-coil+protein+microfibers.">Engineered coiled-coil protein microfibers.</a> Biomacromolecules. 15, 3503-3510 (2014).</li></ol>

而评估纳米材料包括CNPS的潜在毒性作用，有人指出，在长期，CNPS被从初始较为分散的颗粒分布到较大 ​​的聚集形式（ 图2）转化。在某些情况下，这些中生产在细胞培养皿，生物条件下高度聚集的地层，形成了从中央骨料含有“海胆”高度线性突起，令人联想起先前所述铜的6。应当指出的是，这里所示的条件下，CNPS的浓度加入到细胞中是次最大，因此不杀死所有的细胞?...

Copper is a highly reactive metal that in the biological world is essential in some enzyme functions 1,2, but in higher concentrations is potently toxic including in the nanoparticulate form 3,4. Concern over copper toxicity has become more relevant as CNPs and other copper-based nanomaterials are utilized, due to the increased surface area/mass for nanostructures. Thus, even a small mass of copper, in nanoparticle form, could cause local toxicity due to its ability to penetrate the cell and be broken down into reactive forms. Some biological species can complex with and chelate metal ions, and even incorporate them into biological structures as has been described in marine mussels 5. In studying the potential toxic effects of nanomaterials 4, it was discovered that over time, and under biological conditions used for typical cell culturing (37 &#176;C and 5% CO2), stable copper biocomposites could be formed with a high-aspect ratio (linear) structure.
By a process of elimination, the initial discovery of these linear biocomposites, which occurred in complete cell culture media, was simplified to a defined protocol of essential elements needed for the biocomposites to self-assemble. Self-assembly of two types of highly linear biocomposites was discovered to be possible with two starting metal components: 1) CNPs and 2) copper sulfate, with the common biological component being cystine. Although more complex, so called &#8220;urchin&#8221; and &#8220;nanoflower&#8221; type copper-containing structures with nanoscale and microscale features have been previously reported, these were produced under non-biological conditions, such as temperatures of 100 &#176;C or greater 6-8. To our knowledge, synthesis of individual, linear copper-containing nanostructures that are scalable in liquid phase under biological conditions has not been previously described. 
One of the starting materials utilized for synthesis of nanocomposites, namely CNPs, has been reported previously to be very toxic to cells 4. It has recently been reported that after the nanocomposites are formed, these structures are less toxic on a per mass basis than the starting NPs 9. Thus, the synthesis described here may be derived from a biological and biochemical reaction that has utility in stabilizing reactive copper species, both in the sense of transforming the NP form into larger structures and in producing composites less toxic to cells. 
In contrast to many other nanomaterial forms which are known to aggregate or clump upon interaction with biological liquid media 10,11, once formed, the highly linear composites described here avoid aggregation, possibly due to a redistribution of charge which has been previously reported 9. As detailed in the current work, this avoidance of aggregation is convenient for the purposes of working with the structures once formed for at least 3 reasons: 1) composite structures once formed may be concentrated using centrifugation and then easily dispersed again using vortex mixing; 2) formed structures can be decreased in average size by sonication for different periods of time; and 3) the formed linear structures may provide an additional tool for avoiding the recently described &#8220;coffee ring effect&#8221; 12 and thus provide a dopant for creating more evenly distributed coatings of materials, especially those containing spherical particulates.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Mini Vortexer</td>
			<td>VWR (https://us.vwr.com)</td>
			<td>58816-121</td>
			<td></td>
		</tr>
		<tr>
			<td>CO2 Incubator Model # 2425-2</td>
			<td>VWR (https://us.vwr.com)</td>
			<td>Contact vendor</td>
			<td>Current model calalog # 98000-360</td>
		</tr>
		<tr>
			<td>Eppendorf Centrifuge (Refrigerated Microcentrifuge)</td>
			<td>Labnet (http://labnetinternational.com/)</td>
			<td>C2500-R</td>
			<td>Model Prism R</td>
		</tr>
		<tr>
			<td>Cell Culture Centrifuge Model Z323K</td>
			<td>Labnet (http://labnetinternational.com/)</td>
			<td>Contact vendor</td>
			<td>Current model Z206A catalog # C0206-A</td>
		</tr>
		<tr>
			<td>Sonicator (Ultrasonic Cleaner)</td>
			<td>Branson Ultrasonics Corporation (http://www.bransonic.com/)</td>
			<td>1510R-MTH</td>
			<td></td>
		</tr>
		<tr>
			<td>Balance</td>
			<td>Sartorius (http://dataweigh.com)</td>
			<td></td>
			<td>Model CP225D similar model CPA225D</td>
		</tr>
		<tr>
			<td>Olympus IX51 Inverted Light Microscope</td>
			<td>Olympus (http://olympusamerica.com</td>
			<td>Contact vendor</td>
			<td></td>
		</tr>
		<tr>
			<td>Olympus DP71 microscope digital camera</td>
			<td>Olympus (http://olympusamerica.com</td>
			<td>Contact vendor</td>
			<td></td>
		</tr>
		<tr>
			<td>external power supply unit- white light for Olympus microscope</td>
			<td>Olympus (http://olympusamerica.com</td>
			<td>TH4-100</td>
			<td></td>
		</tr>
		<tr>
			<td>10x, 20, and 40x microscope objectives</td>
			<td>Olympus (http://olympusamerica.com</td>
			<td>Contact vendor</td>
			<td></td>
		</tr>
		<tr>
			<td>Scanning Electron Microscope</td>
			<td>Hitachi (http://hitachi-hitec.com/global/em/sem/sem_index.html)</td>
			<td>model S-4800</td>
			<td></td>
		</tr>
		<tr>
			<td>Transmission Electron Microscope</td>
			<td>Zeiss (http://zeiss.com/microscopy/en_de/products.html)</td>
			<td>model Libra 120</td>
			<td></td>
		</tr>
		<tr>
			<td>Table Top Work Station Unidirectional Flow Clean Bench</td>
			<td>Envirco (http://envirco-hvac.com)</td>
			<td>model PNG62675</td>
			<td>Used for sterile cell culture technique</td>
		</tr>
	</tbody>
</table>

generation of scalable, metallic high-aspect ratio nanocomposites in a biological liquid medium

该协议的目的是描述两种新型生物复合材料具有高纵横比结构的合成。该生物复合材料包括铜和胱氨酸，无论是与铜纳米颗粒（CNPS）或硫酸铜贡献的金属部件。合成是在生物条件下（37℃）和24小时后的自组装的复合材料的形式下是液体。一旦形成，这些复合材料是在两种液体介质和干燥形式高度稳定。该复合物由纳米规模向微型范围在长度和从几微米到直径25纳米。能量色散X射线光谱仪（EDX）的场发射扫描电子显微镜表明，硫存在于NP衍生线性结构，而这是从起始CNP材料不存在，从而证实胱氨酸作为硫在最终纳米复合材料的源。在综合这些线性纳米和微观复合材料，STR的长度多种多样uctures形成在合成容器。合成后的液体混合物的超声处理被证明有助于通过用超声处理的时间增加减少的平均长度控制的结构的平均尺寸。因为所形成的结构是高度稳定的，不结块，并形成有在液相，离心也可用于帮助浓缩和分离形成的复合材料。

图1示出的合成步骤，以形成在这项工作中描述的线性生物复合材料的流程图示意图。 CNPS或硫酸铜作为起始原料相结合，与无菌水，以形成2毫克/毫升溶液，该溶液混合并超声处理以提供更混合物，使该铜溶液然后在下面的比例混合用于合成：949份无菌水：50份混合铜：1份胱氨酸原液。实际体积可以根据这些比率来放大或缩小最后合成收率而增加或减少。孵育至少2小时所指示的，线?...

Watch this Scientific Journal Video about Generation of Scalable, Metallic High-Aspect Ratio Nanocomposites in a Biological Liquid Medium at JoVE.com

Generation of Scalable, Metallic High-Aspect Ratio Nanocomposites in a Biological Liquid Medium

在这里，我们提出了一个协议，以合成下生物条件和在液体介质新颖的，高宽比生物复合材料。的生物复合材料从纳米扩展到微米直径和长度。铜纳米粒子（CNPS）和硫酸铜结合胱氨酸是关键部件的合成。

可扩展的，金属的高宽比纳米复合材料在生物液体中产生

The goal of this protocol is to describe the synthesis of two novel biocomposites with high-aspect ratio structures. The biocomposites consist of copper ...

Research

JoVE Journal

Bioengineering

Cellular Lipid Extraction for Targeted Stable Isotope Dilution Liquid Chromatography-Mass Spectrometry Analysis

有针对性的稳定同位素稀释液相色谱 - 质谱法分析细胞脂质提取

The metabolism of fatty acids, such as arachidonic acid (AA) and linoleic acid (LA), results in the formation of oxidized bioactive lipids, including ...

科研

生物工程

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

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

环境控制机械自适应聚合物纳米复合材料的微拉伸测试<em&gt;体外</em&gt;表征

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

In situ TEM of Biological Assemblies in Liquid

In situ TEM of Biological Assemblies in Liquid

就地 液体生物组件的 TEM

Researchers regularly use Transmission Electron Microscopes (TEMs) to examine biological entities and to assess new materials. Here, we describe an ...

Synthesis of Keratin-based Nanofiber for Biomedical Engineering

基于角蛋白纳米纤维生物医学工程合成

Electrospinning, due to its versatility and potential for applications in various fields, is being frequently used to fabricate nanofibers. Production of ...

Alternating Magnetic Field-Responsive Hybrid Gelatin Microgels for Controlled Drug Release

交变磁场敏感的混合明胶微凝胶药物控释

Magnetically-responsive nano/micro-engineered biomaterials that enable a tightly controlled, on-demand drug delivery have been developed as new types of ...

Preparation of Monodomain Liquid Crystal Elastomers and Liquid Crystal Elastomer Nanocomposites

单域液晶的弹性体和液晶橡胶纳米复合材料的制备

LCEs are shape-responsive materials with fully reversible shape change and potential applications in medicine, tissue engineering, artificial muscles, and ...

A Novel Technique for Generating and Observing Chemiluminescence in a Biological Setting

在生物环境的新技术的产生和观测化学发光

Intraoperative imaging techniques have the potential to make surgical interventions safer and more effective; for these reasons, such techniques are ...

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

模块化 DNA 设备的自动机器人液体处理组件

Recent advances in modular DNA assembly techniques have enabled synthetic biologists to test significantly more of the available &#34;design space&#34; ...

Synthesis of Biocompatible Liquid Crystal Elastomer Foams as Cell Scaffolds for 3D Spatial Cell Cultures

生物相容的液晶弹性体泡沫的合成中用作细胞支架的三维空间细胞培养

Here, we present a step-by-step preparation of a 3D, biodegradable, foam-like cell scaffold. These scaffolds were prepared by cross-linking star block ...

A Microcontroller Operated Device for the Generation of Liquid Extracts from Conventional Cigarette Smoke and Electronic Cigarette Aerosol

一种单片机操作装置, 用于生成传统卷烟烟雾和电子烟气溶胶中的液体萃取物

Electronic cigarettes are the most popular tobacco product among middle and high schoolers and are the most popular alternative tobacco product among ...