Este trabajo de investigación se admite como parte del Centro para el nivel atómico Catalizador Design, un centro de investigación de frontera para la energía financiado por el Departamento de Energía de los EE.UU., Oficina de Ciencia, Oficina de Ciencias Básicas de Energía en virtud de Premio Número DE-SC0001058 y también con el apoyo de la Junta de Regents en las subvenciones del premio número LEQSF (2009-14)-EFRC-MATCH y LEDSF-EPS (2012)-OPT-IN-15. Operaciones MRCAT son apoyados por el Departamento de Energía y las instituciones miembros MRCAT. El uso de la Fuente Avanzada de Fotones en ANL es apoyado por el Departamento de Energía de EE.UU., Oficina de Ciencia, Oficina de Ciencias Básicas de Energía, bajo el Contrato No. DE-AC02-06CH11357. El apoyo financiero para JTM se proporciona como parte del Instituto de Atom-eficientes Transformaciones Químicas (IACT), un Centro de Investigación en Energía de la Frontera, financiado por el Departamento de Energía de los EE.UU., Oficina de Ciencia, Oficina de Ciencias Básicas de Energía.

Millifluidics set-up: Compra un chip millifluidic (hecha de polímero de tereftalato de poliéster) de Microplumbers Microsciences LLC, que tiene canales serpenteantes con dimensiones de 2 mm (W) x 0,15 mm (H) x 220 mm (L). Utilice FEP Tubería con dimensiones de 0,25 mm de diámetro, 1/16 en OD, para conectar el chip a la bomba. Utilice dos bombas diferentes para los dos experimentos diferentes. Usar P-Bomba para el primer experimento (nanopartículas de cobre) y el dispositivo millifluidic para el segundo experimento (nanopartículas de oro). Para minimizar el problema de las burbujas de gas dentro de los canales, solución recién preparada NaBH4 se dejó abierta en reposo durante ~ 10-15 minutos antes de bombear en el chip de manera que las burbujas de gas escapen de la solución. Este paso fue seguido por todos nuestros experimentos. 1. Síntesis de Ultra-pequeños agregados de cobre nano (UCNCs) <ol><li> Productos químicos requeridos: Obtener cobre (II) nitrahidrato de TE, borohidruro de sodio, gránulos de hidróxido de sodio y O-[2 - (3-mercaptopropionilamino) etil] glicol-O &#39;metilpolietilenglicol (PM = 5000) [MPEG] y el uso de todos los productos químicos sin purificación adicional. Use agua nanopura (18,2 MΩ-cm) para el experimento. </li><li> Usar P-bomba regulada bajo presión de nitrógeno para el experimento. Prueba de las bombas con agua como disolvente a diferentes presiones antes del experimento que se correlaciona con los caudales correspondientes (ml / h). Enjuague el reactor millifluidic y tubos con agua desionizada antes del inicio del experimento. </li><li> Disolver 174 mg (0,95 mmol) de cobre (II) nitrato y 610 mg (0,122 mmol) de O-[2 - (3-mercaptopropionilamino) etil] glicol-O &#39;metilpolietilenglicol en 28 ml de agua nanopura y mantenerlos en un vial para ser conectado con un canal de entrada </li><li> Mantener otra solución de 111 mg (2,93 mmol) de borohidruro de sodio y 102 mg (2,78 mmol) de hidróxido de sodio en 28 ml (pH ~ 13) en un vial diferente y conectarse con laotro canal de entrada. </li><li> El flujo tanto de las soluciones de forma simultánea dentro del reactor millifluidic a diferentes caudales (que figuran a continuación) y recoger el UCNCs resultantes en la salida en vial de vidrio. Se purga la solución con nitrógeno y almacenarlo bajo nitrógeno. </li><li> Operar las bombas bajo diferentes presiones constantes de 50 mbar (6,81 ml / h), 100 mbar (14,31 ml / h), 200 mbar (32,7 ml / h) y 300 mbar (51,4 ml / h) a temperatura ambiente durante la síntesis de UCNCs a diferentes caudales. </li></ol> Mientras que el procedimiento de síntesis se demostró utilizando la millifluidic puesta a punto con P-bomba, sino que también puede llevarse a cabo utilizando el dispositivo millifluidic de mano de Millifluidica. 2. Tiempo Resuelto En estudios cinéticos in situ en las nanopartículas de oro Formación <ol><li> Productos químicos requeridos: Obtener ácido cloroáurico (HAuCl4 3H 2 O). Meso-2 ,3-dimercaptosuccínico (DMSA) y borohidruro de sodio y #160, y utilizar todos los productos químicos sin purificación adicional. Use agua nanopura (18,2 MΩ-cm) para el experimento. </li><li> El uso de alta precisión, bombas de jeringa, libre de pulsaciones totalmente automatizados fluyan los líquidos dentro del chip. Prueba de las bombas con agua como disolvente en diferentes índices de flujo antes del experimento para optimizar la velocidad de flujo requerida. </li><li> Preparar soluciones estándar de (i) HAuCl 4. 3H 2 O (10 mmol, 118,2 mg/30 ml) y (ii) DMSA (20 mmol, 109,2 mg/30 ml) con 50 mg de hidróxido de sodio (pH 12) en nanopura agua. </li><li> Alimentar a los dos soluciones a través de dos jeringas separadas en el chip millifluidic a una velocidad de flujo constante de 10 ml / h con la bomba automatizada. </li><li> Pareja el chip millifluidic a la línea de haz de sincrotrón utilizando una etapa de metal que tiene acceso a movimiento en direcciones XYZ y recoger los datos XAS en diferentes zonas en el chip como las soluciones se bombearon a través del chip. </li></ol> Mientras que el &lt;em&gt; en análisis in situ procedimiento se demostró utilizando la millifluidic puesta a punto con P-bomba, puede también llevarse a cabo utilizando un dispositivo de millifluidic de mano. 3. Flujo continuo de oro Catálisis Este procedimiento se demostró utilizando un dispositivo millifluidic de mano. <ol><li> Productos químicos requeridos: obtener el ácido cloroáurico (HAuCl 4 3H 2 O.), Meso-2 ,3-dimercaptosuccínico (DMSA), borohidruro de sodio, 4-nitrofenol, 4-aminofenol y el uso de todos los productos químicos sin purificación adicional. Use agua nanopura (18,2 MΩ-cm) para el experimento. </li><li> Preparación del catalizador:. Preparar soluciones patrón de HAuCl4 3H 2 O (10 mmol, 118,2 mg/30 ml), DMSA (20 mmol, 109,2 mg/30 ml) y NaBH4 (10 mmol, 11,34 mg/30 ml) en Nanopure agua. </li><li> Tomar 10 ml cada una de HAuCl4 y soluciones DMSA en dos viales separados y th fluirem en el chip usando el dispositivo millifluidic de mano con un caudal uniforme de 12 ml / h durante 45 min. </li><li> El flujo 10 mmoles NaBH4 dentro del chip a 12 ml / h de caudal durante 15 minutos para reducir el Au (I) en Au (0). </li><li> Finalmente, lavar el chip con agua nanopura durante 30 min a la misma velocidad de flujo antes de realizar los experimentos de catálisis. </li><li> Catálisis de reacción: realizar la reacción de conversión química (reducción) de 4-nitrofenol (4-NP) en 4-aminofenol (4-AP) en el catalizador de oro (preparado anteriormente) recubierto canal millifluidic como se indica a continuación. </li><li> Mezclar 15 ml de solución de 9 x 10 -5 moles de 4-NP con 3,3 ml de 0,65 mol NaBH4 solución para formar iones de 4-nitrofenolato (4-NPI). </li><li> Pasar la solución resultante sobre el catalizador de oro depositadas en el chip a una velocidad de flujo constante de 5 ml / hora de evaluar la actividad catalítica. Analizar los espectros UV-Vis de los productos recogidos dentro de la gama de longitud de onda de 250 a 500 nm para confirmar la conversión de 4-NP. </li><li> Estimar la actividad catalítica de la reacción mediante la obtención de la curva de calibración de 4-NPI. Curva de calibración puede ser adquirida mediante el trazado de la intensidad de absorción observado experimentalmente (I) de 4-NPI a diferentes concentraciones estándar. Las alturas de los picos (a 399 nm) para las curvas de absorción UV-Vis representan la intensidad de absorción valores (I) y de acuerdo a la ley del de Beer Lambert, cualquier cambio en el valor de la altura del pico mostrarían cambio correspondiente en su concentración. Por lo tanto, estimar la actividad catalítica mediante la búsqueda de la diferencia en las concentraciones iniciales y finales de la sustancia reaccionante de la curva de calibración. Por ejemplo, si la altura del pico es 1 unidad (Figura 6) que corresponde a una conversión catalítica de 90% (basado en el gráfico de calibración). </li></ol>

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Todos los autores excepto CSSR Kumar declarar que no tienen intereses financieros en competencia. C. SSR Kumar es el fundador de la empresa Millifluidica LLC.

Los UCNCs fueron formados por la reacción de reducción de nitrato de cobre con borohidruro de sodio en presencia del agente de recubrimiento polimérico O-[2 - (3-mercaptopropionilamino) etil]-O &#39;metilpolietilenglicol (PM = 5000) [MPEG]. La reacción se lleva a cabo dentro del reactor de chips millifluidic a diferentes caudales, tales como 6,8 ml / h, 14,3 ml / h, 32,7 ml / h, y 51,4 ml / h para estudiar el efecto de los caudales en los UCNCs formados. Los tiempos de residencia respectivos para los caudales anteri...

(LOC) dispositivos Lab-on-a-chip para la síntesis química han demostrado ventaja significativa en términos de aumento de la transferencia de masa y calor, control de reacción superior, de alto rendimiento y más seguro ambiente de la operación 1. Estos dispositivos se pueden clasificar en fluidos de chips con sede y dispositivos fluídicos nonchip basado. Entre los fluidos basados ​​en chips, microfluidos está bien investigado y un tema bien cubierto en la literatura 2-5. Sistemas LOC Nonchip basadas utilizan reactores tubulares 6. Convencionalmente, los sistemas de microfluidos se utilizan para un control preciso y la manipulación de fluidos que están geométricamente limitados a escala submilimétrica. Recientemente hemos introducido el concepto de millifluidics basadas en chip, que se pueden utilizar para la manipulación de fluidos en los canales en escala milimétrica (ya sea ancho o profundidad o ambos de los canales son al menos un milímetro de tamaño) 7-9. Además, los chips de millifluidic son relativamente fáciles de fabricar WHIle ofrece un control similar sobre caudales y la manipulación de reactivos. Estos chips también podrían ser operadas a mayores caudales, la creación de tiempos de residencia más pequeñas, por lo tanto, ofrece la posibilidad de ampliación de la síntesis controlada de nanopartículas con distribución de tamaño estrecha. Como ejemplo, recientemente hemos demostrado la síntesis de ultra pequeños nanoclusters de cobre y les caracterizado utilizando en la espectroscopia de absorción de rayos X in situ, así como TEM. Capacidad para obtener pequeñas tiempos de residencia dentro de los canales millifluidic en combinación con el uso de MPEG, que es bidentado muy eficiente PEGilado agente estabilizador para la formación de coloides estables de nanoclusters de cobre 7. Además de la síntesis de los productos químicos y los nanomateriales, los millifluidics podrían ofrecer, debido al mayor volumen y la concentración en el área de la sonda, una plataforma sintético que es más generalizado y eficaz para los estudios cinéticos resueltas en el tiempo y también achieVes una mejor relación señal-ruido que los sistemas de microfluidos 7,10. Se demuestra el uso de chip de millifluidic como un ejemplo para el análisis de resolución temporal del crecimiento de nanoestructuras de oro de la solución utilizando in situ XAS con una resolución de tiempo tan pequeño como 5 mseg 11. También, la mayoría de los reactores de micro desarrollados hasta la fecha para aplicaciones de catálisis se basan en 12,13 de silicio. Su fabricación es caro, además de pequeños volúmenes generados los hace inadecuados para la fabricación a gran escala. Los dos métodos generales para el revestimiento de los canales con nanocatalizadores - químicas y físicas, a menudo denominados como procedimientos de revestimiento de silicio, son actualmente en boga 14,15. Además de la fabricación de micro caro, la obstrucción de los canales hace que la catálisis de micro reactor puede no ser adecuado para la fabricación a gran escala. Aunque microrreactores se han utilizado para la catálisis heterogénea en micro-flujo continuo a través de procesos EARLIer 16-18, la capacidad de controlar la dimensión y morfología de los catalizadores nanoestructurados de oro incrustados dentro de los canales de flujo continuo nunca fue explorado antes. Recientemente hemos desarrollado una tecnología para el recubrimiento de los canales millifluidic con Au catalizadores, después de haber controlado la morfología nano y dimensiones (Figura 5) 11, para llevar a cabo la catálisis de reacciones químicas industrialmente importantes. A modo de ejemplo, hemos demostrado la conversión de 4-nitrofenol en 4-aminofenol catalizada por oro nanoestructurado recubierto dentro de los canales millifluidic. Teniendo en cuenta que un solo chip reactor de millifluidic puede producir índices de flujo de 50-60 ml / h, 7 de alto rendimiento y la síntesis controlada de productos químicos es posible ya sea a través de la operación de flujo continuo o el procesamiento en paralelo. Con el fin de aprovechar las posibilidades que ofrecen las millifluidics, con algunos ejemplos que se describen como antes, también demuestran un fácil de usardispositivo millifluidic que es portátil y tiene los todos los componentes necesarios, como los chips millifluidic, colectores, controladores de flujo, bombas y conexiones eléctricas integrada. Tal dispositivo millifluidic, como se muestra en la Figura 7, que ahora está disponible en la empresa Millifluidica LLC ( <a href="http://www.millifluidica.com" target="_blank">www.millifluidica.com</a> ). El manuscrito también proporciona protocolos utilizando el dispositivo de millifluidic de mano, tal como se describe a continuación, para la síntesis controlada de nanomateriales, análisis resuelta en el tiempo de los mecanismos de reacción y la catálisis de flujo continuo.

<table border="1">
	<tbody>
		<tr>
			<td>Copper (II) nitrate hydrate</td>
			<td>Sigma-Aldrich</td>
			<td>13778-31-9</td>
			<td>99.999% pure</td>
		</tr>
		<tr>
			<td>O-[2-(3-mercaptopropionylamino)ethyl]-O&#8242;-methylpolyethylene glycol</td>
			<td>Sigma-Aldrich</td>
			<td>401916-61-8&#160;</td>
			<td>MW=5,000</td>
		</tr>
		<tr>
			<td>HAuCl4.3H2O (Chloroauric acid)</td>
			<td>Sigma-Aldrich</td>
			<td>27988-77-8&#160;</td>
			<td>99.999% pure</td>
		</tr>
		<tr>
			<td>meso-2,3-dimercaptosuccinic acid (DMSA)</td>
			<td>Sigma-Aldrich</td>
			<td>304-55-2</td>
			<td>~98% pure</td>
		</tr>
		<tr>
			<td>4-Nitrophenol</td>
			<td>Sigma-Aldrich</td>
			<td>100-02-7&#160;</td>
			<td>spectrophotometric grade</td>
		</tr>
		<tr>
			<td>4-Aminophenol</td>
			<td>Sigma-Aldrich</td>
			<td>123-30-8&#160;</td>
			<td>&#62;99% pure (HPLC grade)</td>
		</tr>
		<tr>
			<td>Sodium borohydride</td>
			<td>Sigma-Aldrich</td>
			<td>16940-66-2&#160;</td>
			<td>98% pure</td>
		</tr>
		<tr>
			<td>Sodium hydroxide pellets</td>
			<td>Sigma-Aldrich</td>
			<td>1310-73-2&#160;</td>
			<td>99.99% pure</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>[header]</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>EQUIPMENT</td>
		</tr>
		<tr>
			<td>Millifluidic Chips</td>
			<td>Microplumbers Microsciences LLC</td>
			<td>SDC-01</td>
			<td>Made from polyester terephthalate polymer</td>
		</tr>
		<tr>
			<td>Pressure Pump</td>
			<td>Mitos P-Pump, Dolomite</td>
			<td>3200016</td>
			<td></td>
		</tr>
		<tr>
			<td>Automated Syringe Pump</td>
			<td>Cetoni Automation and Microsystems, GmbH</td>
			<td></td>
			<td>Syringe pump neMESYS</td>
		</tr>
		<tr>
			<td>UV-3600 UV-VIS-NIR Spectrophotometer</td>
			<td>Shimadzu</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Hand-held Millifluidic Device</td>
			<td>Millifluidica</td>
			<td>SCMD-1008</td>
			<td>Figure 7</td>
		</tr>
	</tbody>
</table>

millifluidics for chemical synthesis and time-resolved mechanistic studies

Procedimientos que utilizan dispositivos millifluidic para la síntesis química y estudios sobre el mecanismo de resolución temporal se describen tomando tres ejemplos. En la primera, se describe la síntesis de ultra pequeños nanoclusters de cobre. El segundo ejemplo proporciona su utilidad para la investigación de tiempo resuelto cinética de las reacciones químicas mediante el análisis de la formación de nanopartículas de oro utilizando en la espectroscopia de absorción de rayos X in situ. El último ejemplo demuestra la catálisis de reacciones de flujo continuo dentro del canal millifluidic recubiertos con catalizador nanoestructurado.

Bien dispersado y nanoclusters de cobre de tamaño uniforme con una distribución de tamaño estrecha se obtuvieron utilizando la configuración de chip de millifluidic (Fig. 1a). Los diferentes índices de flujo utilizados para la síntesis no tienen un efecto significativo en el tamaño de los grupos. Sin embargo, con el aumento de la velocidad de flujo, hay una mejora observable en el estrechamiento de la distribución del tamaño. UCNCs con una mejor distribución de tamaño estrecha se obtuvieron en un caudal de 32...

Watch this Scientific Journal Video about Millifluidics for Chemical Synthesis and Time-resolved Mechanistic Studies at JoVE.com

Millifluidics for Chemical Synthesis and Time-resolved Mechanistic Studies

Dispositivos Millifluidic se utilizan para la síntesis controlada de nanomateriales, análisis de resolución temporal de los mecanismos de reacción y la catálisis de flujo continuo.

Millifluidics para la síntesis química y Estudios mecanicistas Tiempo de resolver

Procedures utilizing millifluidic devices for chemical synthesis and time-resolved mechanistic studies are described by taking three examples. In the ...

millifluidics-for-chemical-synthesis-time-resolved-mechanistic

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SCIENTISTS IN ACTION

11.1K vistas.  Louisiana State University.  Dispositivos Millifluidic se utilizan para la síntesis controlada de nanomateriales, análisis de resolución temporal de los mecanismos de reacción y la catálisis de flujo continuo. 

Video: Millifluidics for Chemical Synthesis and Time-resolved Mechanistic Studies

Combinatorial Synthesis of and High-throughput Protein Release from Polymer Film and Nanoparticle Libraries

Polyanhydrides are a class of biomaterials with excellent biocompatibility and drug delivery capabilities. While they have been studied extensively with conventional one-sample-at-a-time synthesis techniques, a more recent high-throughput approach has been developed enabling the synthesis and testing of large libraries of polyanhydrides1. This will facilitate more efficient optimization and design process of these biomaterials for drug and vaccine delivery applications. The method in this work describes the combinatorial synthesis of biodegradable polyanhydride film and nanoparticle libraries and the high-throughput detection of protein release from these libraries. In this robotically operated method (Figure 1), linear actuators and syringe pumps are controlled by LabVIEW, which enables a hands-free automated protocol, eliminating user error. Furthermore, this method enables the rapid fabrication of micro-scale polymer libraries, reducing the batch size while resulting in the creation of multivariant polymer systems. This combinatorial approach to polymer synthesis facilitates the synthesis of up to 15 different polymers in an equivalent amount of time it would take to synthesize one polymer conventionally. In addition, the combinatorial polymer library can be fabricated into blank or protein-loaded geometries including films or nanoparticles upon dissolution of the polymer library in a solvent and precipitation into a non-solvent (for nanoparticles) or by vacuum drying (for films). Upon loading a fluorochrome-conjugated protein into the polymer libraries, protein release kinetics can be assessed at high-throughput using a fluorescence-based detection method (Figures 2 and 3) as described previously1. This combinatorial platform has been validated with conventional methods2 and the polyanhydride film and nanoparticle libraries have been characterized with 1H NMR and FTIR. The libraries have been screened for protein release kinetics, stability and antigenicity; in vitro cellular toxicity, cytokine production, surface marker expression, adhesion, proliferation and differentiation; and in vivo biodistribution and mucoadhesion1-11. The combinatorial method developed herein enables high-throughput polymer synthesis and fabrication of protein-loaded nanoparticle and film libraries, which can, in turn, be screened in vitro and in vivo for optimization of biomaterial performance.

This method describes the combinatorial synthesis of biodegradable polyanhydride film and nanoparticle libraries and the high-throughput detection of protein release from these libraries.

Síntesis combinatoria y de alto rendimiento liberación de proteínas de película de polímero y nanopartículas Bibliotecas

Polyanhydrides are a class of biomaterials with excellent biocompatibility and drug delivery capabilities. While they have been studied extensively with ...

Investigación

Bioingeniería

High-throughput Synthesis of Carbohydrates and Functionalization of Polyanhydride Nanoparticles

Transdisciplinary approaches involving areas such as material design, nanotechnology, chemistry, and immunology have to be utilized to rationally design efficacious vaccines carriers. Nanoparticle-based platforms can prolong the persistence of vaccine antigens, which could improve vaccine immunogenicity1. Several biodegradable polymers have been studied as vaccine delivery vehicles1; in particular, polyanhydride particles have demonstrated the ability to provide sustained release of stable protein antigens and to activate antigen presenting cells and modulate immune responses2-12.
The molecular design of these vaccine carriers needs to integrate the rational selection of polymer properties as well as the incorporation of appropriate targeting agents. High throughput automated fabrication of targeting ligands and functionalized particles is a powerful tool that will enhance the ability to study a wide range of properties and will lead to the design of reproducible vaccine delivery devices.
The addition of targeting ligands capable of being recognized by specific receptors on immune cells has been shown to modulate and tailor immune responses10,11,13 C-type lectin receptors (CLRs) are pattern recognition receptors (PRRs) that recognize carbohydrates present on the surface of pathogens. The stimulation of immune cells via CLRs allows for enhanced internalization of antigen and subsequent presentation for further T cell activation14,15. Therefore, carbohydrate molecules play an important role in the study of immune responses; however, the use of these biomolecules often suffers from the lack of availability of structurally well-defined and pure carbohydrates. An automation platform based on iterative solution-phase reactions can enable rapid and controlled synthesis of these synthetically challenging molecules using significantly lower building block quantities than traditional solid-phase methods16,17.
Herein we report a protocol for the automated solution-phase synthesis of oligosaccharides such as mannose-based targeting ligands with fluorous solid-phase extraction for intermediate purification. After development of automated methods to make the carbohydrate-based targeting agent, we describe methods for their attachment on the surface of polyanhydride nanoparticles employing an automated robotic set up operated by LabVIEW as previously described10. Surface functionalization with carbohydrates has shown efficacy in targeting CLRs10,11 and increasing the throughput of the fabrication method to unearth the complexities associated with a multi-parametric system will be of great value (Figure 1a).

In this article, a high throughput method is presented for the synthesis of oligosaccharides and their attachment to the surface of polyanhydride nanoparticles for further use in targeting specific receptors on antigen presenting cells.

De alto rendimiento de síntesis de carbohidratos y funcionalización de nanopartículas polianhidrido

Transdisciplinary approaches involving areas such as material design, nanotechnology, chemistry, and immunology have to be utilized to rationally design ...

Microfabrication of Nanoporous Gold Patterns for Cell-material Interaction Studies

Nanostructured materials with feature sizes in tens of nanometers have enhanced the performance of several technologies, including fuel cells, biosensors, biomedical device coatings, and drug delivery tools. Nanoporous gold (np-Au), produced by a nano-scale self-assembly process, is a relatively new material that exhibits large effective surface area, high electrical conductivity, and catalytic activity. These properties have made np-Au an attractive material to scientific community. Most studies on np-Au employ macro-scale specimens and focus on fundamental science of the material and its catalytic and sensor applications. The macro-scale specimens limit np-Au's potential in miniaturized systems, including biomedical devices. In order to address these issues, we initially describe two different methods to micropattern np-Au thin films on rigid substrates. The first method employs manually-produced stencil masks for creating millimeter-scale np-Au patterns, while the second method uses lift-off photolithography to pattern sub-millimeter-scale patterns. As the np-Au thin films are obtained by sputter-deposition process, they are compatible with conventional microfabrication techniques, thereby amenable to facile integration into microsystems. These systems include electrically-addressable biosensor platforms that benefit from high effective surface area, electrical conductivity, and gold-thiol-based surface bioconjugation. We describe cell culture, immunostaining, and image processing techniques to quantify np-Au's interaction with mammalian cells, which is an important performance parameter for some biosensors. We expect that the techniques illustrated here will assist the integration of np-Au in platforms at various length-scales and in numerous applications, including biosensors, energy storage systems, and catalysts.

We report on techniques to micropattern nanoporous gold thin films via stencil printing and photolithography, as well as methods to culture cells on the microfabricated patterns. In addition, we describe image analysis methods to characterize morphology of the material and the cultured cells using scanning electron and fluorescence microscopy techniques.

Microfabricación de los patrones de oro Nanoporous de Estudios de interacción célula-materiales

Nanostructured materials with feature sizes in tens of nanometers have enhanced the performance of several technologies, including fuel cells, biosensors, ...

Synthesis of an Intein-mediated Artificial Protein Hydrogel

We present the synthesis of a highly stable protein hydrogel mediated by a split-intein-catalyzed protein trans-splicing reaction. The building blocks of this hydrogel are two protein block-copolymers each containing a subunit of a trimeric protein that serves as a crosslinker and one half of a split intein. A highly hydrophilic random coil is inserted into one of the block-copolymers for water retention. Mixing of the two protein block copolymers triggers an intein trans-splicing reaction, yielding a polypeptide unit with crosslinkers at either end that rapidly self-assembles into a hydrogel. This hydrogel is very stable under both acidic and basic conditions, at temperatures up to 50 &#176;C, and in organic solvents. The hydrogel rapidly reforms after shear-induced rupture. Incorporation of a &#34;docking station peptide&#34; into the hydrogel building block enables convenient incorporation of &#34;docking protein&#34;-tagged target proteins. The hydrogel is compatible with tissue culture growth media, supports the diffusion of 20 kDa molecules, and enables the immobilization of bioactive globular proteins. The application of the intein-mediated protein hydrogel as an organic-solvent-compatible biocatalyst was demonstrated by encapsulating the horseradish peroxidase enzyme and corroborating its activity.

We present the synthesis of a split-intein-mediated protein hydrogel. The building blocks of this hydrogel are two protein copolymers each containing a subunit of a trimeric protein that serves as a crosslinker and one half of a split intein. Mixing of the two protein copolymers triggers an intein trans-splicing reaction, yielding a polypeptide unit that self-assembles into a hydrogel. This hydrogel is highly pH- and temperature-stable, compatible with organic solvents, and easily incorporates functional globular proteins.

Síntesis de una proteína artificial mediada Intein hidrogel

We present the synthesis of a highly stable protein hydrogel mediated by a split-intein-catalyzed protein trans-splicing reaction. The building blocks of ...

Synthesis of Keratin-based Nanofiber for Biomedical Engineering

Electrospinning, due to its versatility and potential for applications in various fields, is being frequently used to fabricate nanofibers. Production of these porous nanofibers is of great interest due to their unique physiochemical properties. Here we elaborate on the fabrication of keratin containing poly (&#949;-caprolactone) (PCL) nanofibers (i.e., PCL/keratin composite fiber). Water soluble keratin was first extracted from human hair and mixed with PCL in different ratios. The blended solution of PCL/keratin was transformed into nanofibrous membranes using a laboratory designed electrospinning set up. Fiber morphology and mechanical properties of the obtained nanofiber were observed and measured using scanning electron microscopy and tensile tester. Furthermore, degradability and chemical properties of the nanofiber were studied by FTIR. SEM images showed uniform surface morphology for PCL/keratin fibers of different compositions. These PCL/keratin fibers also showed excellent mechanical properties such as Young&#39;s modulus and failure point. Fibroblast cells were able to attach and proliferate thus proving good cell viability. Based on the characteristics discussed above, we can strongly argue that the blended nanofibers of natural and synthetic polymers can represent an excellent development of composite materials that can be used for different biomedical applications.

Electrospun nanofibers have a high surface area to weight ratio, excellent mechanical integrity, and support cell growth and proliferation. These nanofibers have a wide range of biomedical applications. Here we fabricate keratin/ PCL nanofibers, using the electrospinning technique, and characterize the fibers for possible applications in tissue engineering.

Síntesis de nanofibras a base de Queratina de Ingeniería Biomédica

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

The Synthesis of RGD-functionalized Hydrogels as a Tool for Therapeutic Applications

The use of polymers as biomaterials has provided significant advantages in therapeutic applications. In particular, the possibility to modify and functionalize polymer chains with compounds that are able to improve biocompatibility, mechanical properties, or cell viability allows the design of novel materials to meet new challenges in the biomedical field. With the polymer functionalization strategies, click chemistry is a powerful tool to improve cell-compatibility and drug delivery properties of polymeric devices. Similarly, the fundamental need of biomedicine to use sterile tools to avoid potential adverse-side effects, such as toxicity or contamination of the biological environment, gives rise to increasing interest in the microwave-assisted strategy.
The combination of click chemistry and the microwave-assisted method is suitable to produce biocompatible hydrogels with desired functionalities and improved performances in biomedical applications. This work aims to synthesize RGD-functionalized hydrogels. RGD (arginylglycylaspartic acid) is a tripeptide that can mimic cell adhesion proteins and bind to cell-surface receptors, creating a hospitable microenvironment for cells within the 3D polymeric network of the hydrogels. RGD functionalization occurs through Huisgen 1,3-dipolar cycloaddition. Some PAA carboxyl groups are modified with an alkyne moiety, whereas RGD is functionalized with azido acid as the terminal residue of the peptide sequence. Finally, both products are used in a copper catalyzed click reaction to permanently link the peptide to PAA. This modified polymer is used with carbomer, agarose and polyethylene glycol (PEG) to synthesize a hydrogel matrix. The 3D structure is formed due to an esterification reaction involving carboxyl groups from PAA and carbomer and hydroxyl groups from agarose and PEG through microwave-assisted polycondensation. The efficiency of the gelation mechanism ensures a high degree of RGD functionalization. In addition, the procedure to load therapeutic compounds or biological tools within this functionalized network is very simple and reproducible.

We present a protocol for the synthesis of RGD-functionalized hydrogels as devices for cell and drug delivery. The procedure involves copper catalyzed alkyne-azide cycloaddition (CuAAC) between alkyne-modified polyacrylic acid (PAA) and a RGD-azide derivative. The hydrogels are formed using microwave-assisted polycondensation and their physicochemical properties are investigated.

La síntesis de hidrogeles funcionalizado-RGD como una herramienta para aplicaciones terapéuticas

The use of polymers as biomaterials has provided significant advantages in therapeutic applications. In particular, the possibility to modify and ...

Polydimethylsiloxane-polycarbonate Microfluidic Devices for Cell Migration Studies Under Perpendicular Chemical and Oxygen Gradients

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under combinations of chemical and oxygen gradients. Both chemical and oxygen gradients can greatly affect cell migration in vivo; however, due to technical limitations, very little research has been performed to investigate their effects in vitro. The device developed in this research takes advantage of a series of serpentine-shaped channels to generate the desired chemical gradients and exploits a spatially confined chemical reaction method for oxygen gradient generation. The directions of the chemical and oxygen gradients are perpendicular to each other to enable straightforward migration result interpretation. In order to efficiently generate the oxygen gradients with minimal chemical consumption, the embedded PC thin film is utilized as a gas diffusion barrier. The developed microfluidic device can be actuated by syringe pumps and placed into a conventional cell incubator during cell migration experiments to allow for setup simplification and optimized cell culture conditions. In cell experiments, we used the device to study migrations of adenocarcinomic human alveolar basal epithelial cells, A549, under combinations of chemokine (stromal cell-derived factor, SDF-1&#945;) and oxygen gradients. The experimental results show that the device can stably generate perpendicular chemokine and oxygen gradients and is compatible with cells. The migration study results indicate that oxygen gradients may play an essential role in guiding cell migration, and cellular behavior under combinations of gradients cannot be predicted from those under single gradients. The device provides a powerful and practical tool for researchers to study interactions between chemical and oxygen gradients in cell culture, which can promote better cell migration studies in more in vivo-like microenvironments.

The control of chemical and oxygen gradients is essential for cell cultures. This paper reports a polydimethylsiloxane-polycarbonate (PDMS-PC) microfluidic device capable of reliably generating combinations of chemical and oxygen gradients for cell migration studies, which can be practically utilized in biological labs without sophisticated instrumentation.

Polidimetilsiloxano-policarbonato celular dispositivos de microfluidos para Estudios de Inmigración en virtud perpendicular Química y gradientes de oxígeno

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under ...

Synthesis of Hydrogels with Antifouling Properties As Membranes for Water Purification

Hydrogels have been widely utilized to enhance the surface hydrophilicity of membranes for water purification, increasing the antifouling properties and thus achieving stable water permeability through membranes over time. Here, we report a facile method to prepare hydrogels based on zwitterions for membrane applications. Freestanding films can be prepared from sulfobetaine methacrylate (SBMA) with a crosslinker of poly(ethylene glycol) diacrylate (PEGDA) via photopolymerization. The hydrogels can also be prepared by impregnation into hydrophobic porous supports to enhance the mechanical strength. These films can be characterized by attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) to determine the degree of conversion of the (meth)acrylate groups, using goniometers for hydrophilicity and differential scanning calorimetry (DSC) for polymer chain dynamics. We also report protocols to determine the water permeability in dead-end filtration systems and the effect of foulants (bovine serum albumin, BSA) on membrane performance.

This paper reports practical methods to prepare hydrogels in freestanding films and impregnated membranes and to characterize their physical properties, including water transport properties.

Síntesis de hidrogeles con propiedades antiincrustantes como membranas de purificación de agua

Hydrogels have been widely utilized to enhance the surface hydrophilicity of membranes for water purification, increasing the antifouling properties and ...

Ammonia Synthesis at Low Pressure

Ammonia can be synthesized at low pressure by the use of an ammonia selective absorbent. The process can be driven with wind energy, available locally in areas requiring ammonia for synthetic fertilizer. Such wind energy is often called &#34;stranded,&#34; because it is only available far from population centers where it can be directly used.
In the proposed low pressure process, nitrogen is made from air using pressure swing absorption, and hydrogen is produced by electrolysis of water. While these gases can react at approximately 400 &#176;C in the presence of a promoted conventional catalyst, the conversion is often limited by the reverse reaction, which makes this reaction only feasible at high pressures. This limitation can be removed by absorption on an ammine-like calcium or magnesium chloride. Such alkaline metal halides can effectively remove ammonia, thus suppressing the equilibrium constraints of the reaction. In the proposed absorption-enhanced ammonia synthesis process, the rate of reaction may then be controlled not by the chemical kinetics nor the absorption rates, but by the rate of the recycle of unreacted gases. The results compare favorably with ammonia made from a conventional small scale Haber-Bosch process.

Ammonia can be synthesized at low pressure by using a conventional catalyst and an ammonia selective absorbent.

Síntesis del amoníaco a baja presión

Ammonia can be synthesized at low pressure by the use of an ammonia selective absorbent. The process can be driven with wind energy, available locally in ...

Synthesis of Functionalized 10-nm Polymer-coated Gold Particles for Endothelium Targeting and Drug Delivery

Gold nanoparticles (AuNPs) have been used extensively in medical research due to their size, biocompatibility, and modifiable surface. Specific targeting and drug delivery are some of the applications of these AuNPs, but endothelial extracellular matrices&#39; defensive properties hamper particle uptake. To address this issue, we describe a synthesis method for ultrasmall gold nanoparticles to improve vascular delivery, with customizable functional groups and polymer lengths for further adjustments. The protocol yields 2.5 nm AuNPs that are capped with tetrakis(hydroxymethyl)phosphonium chloride (THPC). The replacement of THPC with hetero-functional polyethylene glycol (PEG) on the surface of the AuNP increases the hydrodynamic radius to 10.5 nm while providing various functional groups on the surface. The last part of the protocol includes an optional addition of a fluorophore to allow the AuNPs to be visualized under fluorescence to track nanoparticle uptake. Dialysis and lyophilization were used to purify and isolate the AuNPs. These fluorescent nanoparticles can be visualized in both in vitro and in vivo experiments due to the biocompatible PEG coating and fluorescent probes. Additionally, the size range of these nanoparticles render them an ideal candidate for probing the glycocalyx without disrupting normal vasculature function, which may lead to improved delivery and therapeutics.

We describe a method of synthesizing biocompatible 10-nm gold nanoparticles, functionalized by coating poly-ethylene glycol onto the surface. These particles can be used in vitro and in vivo for delivering therapeutics to nanoscale cellular and extracellular spaces that are difficult to access with conventional nanoparticle sizes.

Síntesis de funcionalizados 10 nm recubierto de polímero partículas de oro para apuntar el endotelio y el suministro de medicamentos

Gold nanoparticles (AuNPs) have been used extensively in medical research due to their size, biocompatibility, and modifiable surface. Specific targeting ...