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. Planejamento de Experimentos <ol><li> Determinar o volume de nanocompósitos de cobre necessário para a síntese. Nessa base, escolha um número de frascos de pequeno volume (25 cm2), ou frascos maiores, como indicado a seguir na preparação de materiais. </li><li> Para esta síntese, utilizar uma temperatura de 37 ° C incubadora com 5% de CO 2 e, pelo menos, 40% de humidade. Assegure-se que uma tal incubadora está disponível e que não será perturbado repetidamente ao longo do período ...

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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. 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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. 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Ao avaliar os potenciais efeitos tóxicos de nanomateriais incluindo CNPs, observou-se que, a longo prazo, CNPs foram transformados a partir de uma distribuição de partículas mais inicialmente dispersa para uma forma maior, agregados (Figura 2). Em alguns casos, essas formações altamente agregados que foram produzidos na placa de cultura de células, sob condições biológicas, formado projeções altamente linear a partir do centro de agregado, que relembram a cobre anteriormente descrito contend...

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

O objetivo deste protocolo é descrever a síntese de duas novas biocompósitos com estruturas de relação de aspecto de alta. Os biocompósitos composto de cobre e cistina, quer com nanopartículas de cobre (CNPS) ou sulfato de cobre contribuindo o componente metálico. A síntese é levada a cabo no estado líquido sob condições biológicas (37 ° C) e sob a forma de compósitos auto-montada após 24 h. Uma vez formados, estes compostos são altamente estáveis ​​em ambos os meios líquidos e em uma forma seca. Os compósitos escala nano a partir do micro- gama de comprimento, e de alguns microns a 25 nm de diâmetro. A microscopia electrónica de varrimento com emissão de campo espectroscopia de energia dispersiva de raios-X (EDX), demonstrou que o enxofre estava presente nas estruturas lineares NP-derivados, que não foi encontrada a partir do material de partida CNP, confirmando assim cistina como a fonte de enxofre nos nanocompósitos finais . Durante a síntese destes nano e micro-compósitos lineares, uma variada gama de comprimentos de structures é formada no reactor de síntese. A sonicação da mistura líquida após a síntese foi demonstrado para ajudar a controlar a dimensão média das estruturas, diminuindo a duração média com o aumento do tempo de sonicação. Uma vez que as estruturas formadas são altamente estáveis, não se aglomeram, e são formadas na fase líquida, centrifugação, também podem ser utilizados para auxiliar na concentração e segregar compósitos formados.

A Figura 1 mostra um fluxograma esquemático dos passos de síntese, para formar os biocompósitos lineares descritas neste trabalho. CNPs ou sulfato de cobre como materiais de partida são combinados com água esterilizada para formar uma solução / ml 2 mg, esta solução é misturada e sonicada para fornecer uma mistura uniforme, e esta solução de cobre é, em seguida, misturados na seguinte proporção para a síntese de: 949 partes estéril água: 50 partes de mistura de cobre: ​​1 parte de ...

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

Aqui apresentamos um protocolo para sintetizar novos, de alta relação de aspecto biocompósitos condições biológicas e em meio líquido. Os biocompósitos escala de nanómetros a micrómetros de diâmetro e de comprimento, respectivamente. Nanopartículas de cobre (CNPS) e sulfato de cobre combinados com a cistina são os componentes chave para a síntese.

Geração de escalável e de alto-Aspect Metallic nanocompósitos índices, em um Biológico Líquido Médio

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

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