This research was supported by the Technology Innovation Program (Contract grant No. 0037915, WPM Biomedical Materials-Implant Materials) and Industrial Strategic Technology Development Program (Contract grant No. 10045329, Development of customized implant with porous structure for bone replacement), funded by the Ministry of Trade, industry &#38; Energy (MI, Korea), and BK21 PLUS SNU Materials Division for Educating Creative Global Leaders (Contract grant No. 21A20131912052).

1. Herstellung von porösen Metallgerüste <ol><li> Vorbereitung Ti-Camphen Schlämme durch Mischen von kommerziell erhältlichen Ti-Pulver, Camphen und KD-4 nach dem Abwiegen der geeigneten Mengen von Materialien, wie in Tabelle 1 für poröse Ti Gerüste mit vier anfänglichen Porositäten (40, 50, 60 und 70) beschrieben. Gießen der Aufschlämmungen in 500 ml-Polyethylen (PE-Flaschen) und drehen die Flaschen bei 55 ° C für 30 min in einer Kugelmühle Ofen bei 30 Umdrehungen pro Minute. </li><li> Gießen der Schlämme aus den PE-Flaschen in zylindrische Aluminium (Al) Formen mit einem Durchmesser von 60 mm und einer Höhe von 60 mm. Siegel Jede Al Form mit dem entsprechenden Al Deckglas, und drehen Sie die Formen in einer Kugelmühle Ofen bei einer Geschwindigkeit von 30 Umdrehungen pro Minute bei 55 ° C für 10 min. <ol><li> Anschließend senken die Temperatur der Kugelmühle Ofen auf 44 ° C und kontinuierliches Drehen der Formen bei einer Geschwindigkeit von 30 Umdrehungen pro Minute bei einer konstanten Temperatur von 44 ° C für 12 Std. </li></ol></li><li> Nehmen Sie die Form aus dem KugelMühle Backofen, nachdem zusätzlich Drehen der Formen bei RT für 1 h für den Kühlprozess. Entfernen Sie die erstarrten Titan / Camphen Grünkörper aus dem Al Form unter Verwendung eines Al-Kolben. </li><li> Setzen Sie den erstarrten Titan / Camphen Grünkörper in einem Gummisack mit der Hand und der Gummisack vollständig abzudichten durch das Binden der Öffnung des Beutels mit einer Schnur. Legen Sie das Gummisack im Wassertank eines kaltisostatisches Pressen (CIP) Maschine und wenden einen isostatischen Druck von 200 MPa für 10 min. Entfernen Sie die Druckgrünkörper aus der Gummisack. </li><li> Übertragen Sie die Ti-Camphen Grünkörper auf einen Aluminiumoxid-Tiegel mit der Hand und legen Sie den Tiegel im Gefriertrockner Maschine. Gefrierzutrocknen des Grünkörpers, um das Camphen Phase in der Grünkörpers bei sublimieren - 40 ° C für 24 Stunden. </li><li> Anschließend schließen Sie den Tiegel mit einem Aluminiumoxiddeckglas und legen Sie die geschlossenen Tiegel in einem Vakuumofen (unter 10 -6 Torr) bei RT. Dann wird die Temperatur des Ofens auf 1300 ° C bei einer Erwärmungs raß von 5 ° C / min und halten Sie die Temperatur bei 1300 ° C für 2 Stunden. </li><li> Nach der Wärmebehandlung, halten den gesinterten porösen Ti in dem Ofen für 6-7 Stunden, bis der Ofen voll auf RT abgekühlt. Anmerkung: Während der 6 Stunden der Abkühlung ist die mittlere Abkühlungsgeschwindigkeit des Ofens oberhalb von 400 ° C ~ 15 ° C / min und die durchschnittliche Abkühlungsgeschwindigkeit des Ofens unter 400 ° C ~ 2 ° C / min. </li><li> Falls erforderlich, schneiden den Block aus gesintertem porösem Ti in scheibenförmige Proben mit einem Durchmesser von 16 mm durch elektrische Entladungsbearbeitung (EDM). 27 Hinweis: Je nach der Größe der Al Schimmelpilze, die Größe des gesinterten porösen Ti muss durch den Bearbeitungsprozess (2A) verändert werden. </li><li> Einen Glasbecher mit den porösen Ti-Proben in einem Autoklaven sterilisiert und die Proben bei 121 ° C für 15 min. Entfernen der Proben aus dem Autoklaven. Die poröse Ti-Proben mit destilliertem Wasser zweimal zweimal und dann waschen mit 70% Ethanol.Schließlich verlassen die poröse Ti in eine Petrischale und an der Luft trocknen die Proben bei RT auf einem Sterilbank unter UV-Licht. </li></ol> 2. Tauchbeschichtung von Gerüsten mit bioaktiven Mitteln <ol><li> Verdünnen des kommerziellen Green Fluorescence Protein (GFP) von 1 mg / ml bis 100 ug / ml in einer Sterilwerkbank, indem 1 ml von GFP mit 9 ml Dulbeccos phosphatgepufferter Salzlösung (DPBS, pH 7,4) Lösung in einem 10 ml-sterilisiert Polystyrol (PS) Rohr, wie in Tabelle 1 angegeben. </li><li> Tauchen Sie die sterilisiert dicht oder porös Ti in 10 ml verdünnt GFP-Lösung (100 ug / ml), indem Sie die Ti-Proben in das PS-Rohr mit dem GFP-Lösung bei RT und Unterbringung an einem Sterilbank. </li><li> Setzen Sie den PS Rohr im Vakuumtrockenschrank und evakuieren Exsikkator für 10 Minuten, um sicherzustellen, das GFP-Lösung effektiver dringt in die Poren des porösen Ti. </li><li> Entfernen Sie die porösem Titan aus der PS Rohr mit einer Pinzette. Legen Sie das GFP-beschichtete poröse Ti in eine 10 cm Durchmesser Petri Gericht und an der Luft trocknen O / N bei RT auf einem Sterilbank. </li><li> Spülen Sie die poröse Ti zweimal mit 10 ml von Dulbeccos phosphatgepufferter Salzlösung (DPBS) in einem Becherglas, und verschieben Sie die poröse Ti zu einem Durchmesser von 10 cm Petrischale mit einer Pinzette und an der Luft trocknen bei Raumtemperatur auf eine saubere Bank. </li></ol> 3. Verdichtung von porösen Gerüsten <ol><li> Legen Sie die GFP-beschichtete poröse Ti Proben mit verschiedenen Höhen in einem zylindrischen Stahlform, und legen Sie eine Reihe von Schlägen in die oberen und unteren Löcher der Stahlform (3A). </li><li> Komprimieren Sie die poröse Ti innerhalb des Stahlformanordnung bei RT in z-Richtung der Probe (3A) mit einer Pressmaschine bei mittleren Belastungsraten von 0,05 ~ 0,1 s -1 gegen die in Tabelle 2 gezeigten vorbestimmten angewendet Stämme. Halten Sie die Druck 1 min vor der Entladung. </li><li> Entfernen Sie die verdichtete Ti Proben aus der Stahlform. Zweimaliges Waschen der verdichteten Proben mit 10 ml DPBSin einem Becherglas und der Luft trocknen O / N bei RT auf einem Sterilbank. </li></ol> 4. Lassen Test von GFP beschichtet Gerüste <ol><li> Tauchen drei Arten von Proben (GFP beschichteten dichten Ti (nach den Schritten 2), GFP-beschichtete poröse Ti (nach den Schritten 1 und 2) und GFP-beschichtete verdichtete poröse Ti (nach den Schritten 1-3)) in 5 ml DPBS (pH 7,4) Lösung in einem 10 ml sterilisiertem PS Rohr auf eine saubere Bank enthielt bei 37 ° C. </li><li> Absaugen aus ganze DPBS-Lösung von jedem PS Rohr mit dem GFP-beschichteten Probe und füllt mit einer neuen 5 ml DPBS-Lösung (pH 7,4) unter Verwendung einer Pipette gemäß den vorbestimmten Zeiten von 1, 2, 3, 5, 8, 12, 15, 22 und 29 Tage nach der Immersion. </li><li> Nehmen Sie die Fluoreszenzbilder der GFP-beschichteten Proben vor dem Eintauchen (Tag 0) und nach 22 Tage-Tauch mit konfokalen Laser-Scanning-Spektroskopie (CLSM). </li><li> Messung der Fluoreszenzsignalintensität der GFP-Freigabe in 1 ml Lösung von insgesamt 5 ml DPBS-Lösung unter Verwendung von jeder PS Rohr in 4.2 gezeichnetUV-spektroskopisch bei einer Wellenlänge von 215 nm. Den Intensitätswert umzuwandeln in die Konzentration des GFP-Lösung unter Verwendung der Standardkurve. Anmerkung: Vor der Messung wird die Ziehung der Standardkurve für GFP Lösung durch Messen der Fluoreszenzsignalintensität der GFP-Lösung in einem Konzentrationsbereich von 0 ng / ml - 10 ug / ml. </li></ol> 5. Herstellung von Graded Porous Ti Gerüste <ol><li> Erzeugen einen Block des gesinterten porösen Ti durch Wiederholen von Schritt 1.1 zu Schritt 1.7. </li><li> Maschine der gesinterte poröse Ti Block entsprechend der vorbestimmten Struktur-Designs (zB 5a und 5d), die durch EDM. </li><li> Platzieren der bearbeiteten Ti Proben mit Höhenverteilung in einer Stahlform, wo der Durchmesser des porösen Ti ~ 0,1 mm kleiner als der Durchmesser der Düse und legen eine Reihe von Schlägen in die oberen und unteren Löcher der Stahlform. </li><li> Führen Sie die Schritte 3.2 und 3.3. </li></ol> 6. Porosität Measurement Ti Gerüste <ol><li> Messen Sie die Masse (m n) des Ti Gerüsten. </li><li> Berechnung des scheinbaren Volumens (V n) durch Messen der Länge, Breite und Höhe des Ti Gerüsten. </li><li> Berechnen der Porosität unter Verwendung der folgenden Gleichung: <img alt="Gleichung 1" src="/files/ftp_upload/53279/53279eq1.jpg" /> wobei P die Gesamtporosität Prozentsatz ρ Ti beträgt die theoretische Dichte des Titans und m S / V S die gemessene Dichte der Probe. Anmerkung: Die Porosität Ti Proben können direkt von microCT Bildern abgerufen werden, nachdem microCT Bildgebung wird unter Verwendung eines Mikrocomputertomographen durchgeführt wird. </li></ol>

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The authors declare that they have no competing financial interests.

Während Biometall Systeme sind weit verbreitet für biomedizinische Anwendungen verwendet, insbesondere als tragende Materialien, hohe Steifigkeit und geringe Bioaktivität von Metallen haben große Herausforderungen betrachtet. In dieser Studie haben wir die Herstellungsverfahren für eine neue Metall-System, einer verdichteten porösen Metallgerüst, das biomimetische mechanischen Eigenschaften sowie bioaktive Oberfläche mit einer nachhaltigen Freisetzungsverhalten hat. Die Hauptvorteile der Herstellungsverfahren sc...

Während metallische Biomaterialien wurden weithin als tragende Implantate und interne Fixationsvorrichtungen, aufgrund ihrer ausgezeichneten mechanischen Festigkeit und Widerstandskraft, 1-3 verwendet sie beinhalten zwei kritischen Herausforderungen: 1) mechanische Fehlanpassung, weil Metalle sind viel steifer als biologisches Gewebe, was zu unerwünschten Schäden zu den umliegenden Geweben und 2) geringe Bioaktivität, die oft zu einer schlechten Oberfläche mit biologischen Geweben, oft provozieren Fremdkörperreaktionen (zB Entzündung oder Thrombose). 4-6 porösen Metallgerüste vorgeschlagen worden, um das Einwachsen von Knochen in den Strukturen fördern, die Verbesserung . Knochen-Implantat-Kontakt, während die Spannungsabschirmung Effekte werden wegen ihrer geringeren Steifigkeit drückt 7-9 Außerdem können verschiedene Oberflächenmodifikationen wurden angewandt, um die biologischen Aktivitäten von metallischen Implantaten zu verbessern; Solche Modifikationen schließen Beschichtung der Metalloberfläche mit bioaktiven Molekülen (zB Wachstums facToren) oder Arzneimitteln (zB, Vancomycin, Tetracyclin). 10-12 jedoch Probleme wie reduzierte mechanische Eigenschaften der porösen Metallgerüste, verminderte Steifigkeit und die schnelle Freisetzung des biologisch aktiven Überzugsschichten ungelöst bleiben. 13-16 Insbesondere Titan (Ti) und Ti-Legierungen sind eine der beliebtesten Biometall Systeme wegen ihrer ausgezeichneten mechanischen Eigenschaften, chemische Stabilität und gute Biokompatibilität. 13,17-19 Die schaumförmigen Anwendungen wurden zog auch zunehmendes Interesse, weil der 3D- poröse Netzwerke fördern das Einwachsen von Knochen neben knochenartige mechanische Eigenschaften. 20-22 wurden Anstrengungen unternommen, um die mechanischen Eigenschaften durch die Entwicklung neuer Fertigungsverfahren, einschließlich Replikation polymerer Schwamm, Sintermetallpartikeln, Rapid Prototyping (RP) Verfahren zu verbessern, und Platzhalter Verfahren, um die verschiedenen Eigenschaften der Poren zu kontrollieren (zB Porenanteil,Form, Größe, Verteilung und Konnektivität) und Materialeigenschaften (zB metallische Phase und Verunreinigung). 23-25 ​​Vor kurzem hat die Gefriert Gießen von Wasser basierende Metallschlamm beträchtliche Aufmerksamkeit mechanisch verstärkten Ti bildet mit gut ausgerichteten Poren produzieren gewonnen Strukturen durch Ausnutzung der unidirektionalen Eis Dendritenwachstum während der Erstarrung; jedoch Sauerstoffkontamination durch Kontakt von Metallpulvern mit Wasser verursacht erfordert besondere Sorgfalt, um die Versprödung des Ti Gerüsten zu minimieren. 14,15 Daher haben wir einen neuen Ansatz zur Herstellung von bioaktiven und mechanisch abstimmbaren poröse Ti Gerüsten entwickelt. 25. Die Gerüste anfänglich porösen Strukturen mit einer Porosität von mehr als 50%. Die hergestellten porösen Gerüste wurden mit bioaktiven Molekülen beschichtet und dann komprimiert unter Verwendung einer mechanischen Presse, in dem die Endporosität, mechanische Eigenschaften und Arzneimittelfreisetzungsverhaltens wurden vom Anmelder gesteuerteed-Stamm. Die verdichteten porösen Ti Implantate geringer Porosität mit guter Festigkeit trotz der geringen Steifigkeit vergleichbar mit der Knochen (3-20 GPa). 2 Weil der Beschichtungsschicht gezeigt, wurde die Bioaktivität des verdichteten porösen Ti deutlich verbessert. Darüber hinaus kann aufgrund der einzigartigen Flachporenstrukturen durch das Verdichtungsverfahren induziert wurden die beschichteten biologisch aktiven Moleküle gesehen, allmählich vom Gerüst gelöst werden, die Aufrechterhaltung ihrer Wirksamkeit über einen längeren Zeitraum. In dieser Studie haben wir unsere etablierten Methode, um verdichtete poröse Ti Gerüsten für die mögliche Verwendung in biomedizinischen Anwendungen herzustellen. Das Protokoll enthält dynamische Gefrierguss mit Metall Schlämmen und Verdichtung der porösen Gerüsten. Erstens, um poröse Ti Gerüsten mit guter Duktilität der dynamische gefrier Gießverfahren eingeführt wurde, wie in 1A gezeigt, herzustellen. Ti-Pulver wurde in flüssigem Camphen dispergiert sind; Dann wird durch Senken der Temperatur,die flüssige Phase verfestigt, was zur Phasentrennung zwischen dem Ti-Pulver Netzwerk und feste Camphen Kristallen. Anschließend wurde die verfestigte Ti-Camphen Grünkörper gesintert, in dem Ti-Pulver wurden mit kontinuierlicher Ti Streben kondensiert und das Camphen Phase wurde vollständig entfernt, um eine poröse Struktur zu erhalten. Die Beschichtungs- und Verdichtungsverfahren mit den erhaltenen porösen Gerüsten wurde eingesetzt, Variieren der Verdichtungsgrad und erste Porosität. Die Beschichtungsschicht und ihre Freisetzungsverhaltens wurden visualisiert und quantifiziert unter Verwendung des grün fluoreszierenden Proteins (GFP) beschichteten porösen Ti mit oder ohne Verdichtung im Vergleich zu dem GFP-beschichteten dichten Ti. Schließlich wurden funktionsgradierter Ti Gerüsten, die zwei verschiedene poröse Strukturen vorgeschlagen und durch Variieren der Verdichtungsgrad der inneren und äußeren Teile der porösen Gerüsten demonstriert.

<table>
	<tbody>
		<tr>
			<td>Titanium powder</td>
			<td>Alfa Aesar</td>
			<td>#42624</td>
			<td>-325 mesh, 99.5% (metals basis)</td>
		</tr>
		<tr>
			<td>Camphene</td>
			<td>SigmaAldrich</td>
			<td>#456055</td>
			<td>95%, C10H16</td>
		</tr>
		<tr>
			<td>KD-4</td>
			<td>Croda</td>
			<td>&#173;</td>
			<td>Hypermer, polymeric dispersant</td>
		</tr>
		<tr>
			<td>Phosphate Buffer Solution (PBS)</td>
			<td>Welgene</td>
			<td>ML 008-01</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Green Fluorescent Protein (GFP)</td>
			<td>Genoss Co.</td>
			<td>-</td>
			<td>&#62;98% purity, 1mg/ml</td>
		</tr>
		<tr>
			<td>Ball mill oven</td>
			<td>SAMHENUG ENERGY</td>
			<td>SH-BDO150</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Freeze dryer</td>
			<td>Ilshin Lab.</td>
			<td>PVTFD50A</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Cold isostatic pressing (CIP) machine</td>
			<td>SONGWON SYSTEMS</td>
			<td>CIP 42260</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Vaccum furnace</td>
			<td>JEONG MIN INDUSTRIAL</td>
			<td>JM-HP20</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>electical chaege machine</td>
			<td>FANUC robocut</td>
			<td>0iB</td>
			<td>External use</td>
		</tr>
		<tr>
			<td>Press machine</td>
			<td>CG&#38;S</td>
			<td>AJP-200</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Confocal laser scanning spectroscopy (CLSM)</td>
			<td>Olympus</td>
			<td>FluoView FV1000</td>
			<td>External use</td>
		</tr>
	</tbody>
</table>

fabrication of mechanically tunable and bioactive metal scaffolds for biomedical applications

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness and low bioactivity of metals. In this study, we have developed a new method towards fabricating a new type of bioactive and mechanically reliable porous metal scaffolds-densified porous Ti scaffolds. The method consists of two fabrication processes, 1) the fabrication of porous Ti scaffolds by dynamic freeze casting, and 2) coating and densification of the porous scaffolds. The dynamic freeze casting method to fabricate porous Ti scaffolds allowed the densification of porous scaffolds by minimizing the chemical contamination and structural defects. The densification process is distinctive for three reasons. First, the densification process is simple, because it requires a control of only one parameter (degree of densification). Second, it is effective, as it achieves mechanical enhancement and sustainable release of biomolecules from porous scaffolds. Third, it has broad applications, as it is also applicable to the fabrication of functionally graded porous scaffolds by spatially varied strain during densification.

Der Herstellungsprozess, um poröse Ti Gerüsten verwendet wird in 1A dargestellt. Ti-Pulver gehalten wird homogen in Camphen durch kontinuierliche Drehung des Behälters bei 44 ° C 12 h lang dispergiert und während flüssige Camphen vollständig verfestigt ist, irgendwelche Sedimente des relativ schweren Ti Pulvers minimiert werden. Als Ergebnis wurde die homogene Ti-Camphen Grünkörpers unter Verwendung des dynamischen gefrierGussVerfahren hergestellt, wie in 1B gezeigt,</s...

Watch this Scientific Journal Video about Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications at JoVE.com

Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications

Bioactive and mechanically reliable metal scaffolds have been fabricated through a method which consists of two processes, dynamic freeze casting for the fabrication of porous Ti, and coating and densification of the Ti scaffolds. The densification process is simple, effective and applicable to the fabrication of functionally graded scaffolds.

Herstellung von mechanisch Tunable und Bioactive Metallgerüste für biomedizinische Anwendungen

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness ...

fabrication-mechanically-tunable-bioactive-metal-scaffolds-for

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Mechanical Engineering

Bioengineering

Unit 1

Transformations of Stress and Strain

SCIENTISTS IN ACTION

10.6K Aufrufe.  Korea Institute of Industrial Technology. Bioactive and mechanically reliable metal scaffolds have been fabricated through a method which consists of two processes, dynamic freeze casting for the fabrication of porous Ti, and coating and densification of the Ti scaffolds. The densification process is simple, effective and applicable to the fabrication of functionally graded scaffolds.

Serie: Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications

Microfabricated Platforms for Mechanically Dynamic Cell Culture

The ability to systematically probe in vitro cellular response to combinations of mechanobiological stimuli for tissue engineering, drug discovery or fundamental cell biology studies is limited by current bioreactor technologies, which cannot simultaneously apply a variety of mechanical stimuli to cultured cells. In order to address this issue, we have developed a series of microfabricated platforms designed to screen for the effects of mechanical stimuli in a high-throughput format. In this protocol, we demonstrate the fabrication of a microactuator array of vertically displaced posts on which the technology is based, and further demonstrate how this base technology can be modified to conduct high-throughput mechanically dynamic cell culture in both two-dimensional and three-dimensional culture paradigms.

In this protocol, we demonstrate the fabrication of a microactuator array of vertically displaced posts on which the technology is based, and how this base technology can be modified to conduct high-throughput mechanically dynamic cell culture in both two-dimensional and three-dimensional culture paradigms.

Mikrofabrizierten Plattformen für Mechanisch Dynamische Cell Culture

The ability to systematically probe in vitro cellular response to combinations of mechanobiological stimuli for tissue engineering, drug discovery or ...

Forschung

Biotechnik

Preparation of 3D Fibrin Scaffolds for Stem Cell Culture Applications

Stem cells are found in naturally occurring 3D microenvironments in vivo, which are often referred to as the stem cell niche 1. Culturing stem cells inside of 3D biomaterial scaffolds provides a way to accurately mimic these microenvironments, providing an advantage over traditional 2D culture methods using polystyrene as well as a method for engineering replacement tissues 2. While 2D tissue culture polystrene has been used for the majority of cell culture experiments, 3D biomaterial scaffolds can more closely replicate the microenvironments found in vivo by enabling more accurate establishment of cell polarity in the environment and possessing biochemical and mechanical properties similar to soft tissue.3 A variety of naturally derived and synthetic biomaterial scaffolds have been investigated as 3D environments for supporting stem cell growth. While synthetic scaffolds can be synthesized to have a greater range of mechanical and chemical properties and often have greater reproducibility, natural biomaterials are often composed of proteins and polysaccharides found in the extracelluar matrix and as a result contain binding sites for cell adhesion and readily support cell culture. Fibrin scaffolds, produced by polymerizing the protein fibrinogen obtained from plasma, have been widely investigated for a variety of tissue engineering applications both in vitro and in vivo 4. Such scaffolds can be modified using a variety of methods to incorporate controlled release systems for delivering therapeutic factors 5. Previous work has shown that such scaffolds can be used to successfully culture embryonic stem cells and this scaffold-based culture system can be used to screen the effects of various growth factors on the differentiation of the stem cells seeded inside 6,7.
 This protocol details the process of polymerizing fibrin scaffolds from fibrinogen solutions using the enzymatic activity of thrombin. The process takes 2 days to complete, including an overnight dialysis step for the fibrinogen solution to remove citrates that inhibit polymerization. These detailed methods rely on fibrinogen concentrations determined to be optimal for embryonic and induced pluripotent stem cell culture. Other groups have further investigated fibrin scaffolds for a wide range of cell types and applications - demonstrating the versatility of this approach 8-12.

This work details the preparation of 3D fibrin scaffolds for culturing and differentiating plutipotent stem cells. Such scaffolds can be used to screen the effects of various biological compounds on stem cell behavior as well as modified to contain drug delivery systems.

Erstellung von 3D-Fibrin-Gerüste für Stem Cell Culture Anwendungen

Stem cells are found in naturally occurring 3D microenvironments in vivo, which are often referred to as the stem cell niche 1. Culturing stem cells ...

Designing Silk-silk Protein Alloy Materials for Biomedical Applications

Fibrous proteins display different sequences and structures that have been used for various applications in biomedical fields such as biosensors, nanomedicine, tissue regeneration, and drug delivery. Designing materials based on the molecular-scale interactions between these proteins will help generate new multifunctional protein alloy biomaterials with tunable properties. Such alloy material systems also provide advantages in comparison to traditional synthetic polymers due to the materials biodegradability, biocompatibility, and tenability in the body. This article used the protein blends of wild tussah silk (Antheraea pernyi) and domestic mulberry silk (Bombyx mori) as an example to provide useful protocols regarding these topics, including how to predict protein-protein interactions by computational methods, how to produce protein alloy solutions, how to verify alloy systems by thermal analysis, and how to fabricate variable alloy materials including optical materials with diffraction gratings, electric materials with circuits coatings, and pharmaceutical materials for drug release and delivery. These methods can provide important information for designing the next generation multifunctional biomaterials based on different protein alloys.

Blending is an efficient approach to generate biomaterials with a broad range of properties and combined features. By predicting the molecular interactions between different natural silk proteins, new silk-silk protein alloy platforms with tunable mechanical resiliency, electrical response, optical transparency, chemical processability, biodegradability, or thermal stability can be designed.

Gestaltung Silk-Seidenprotein-Legierungen für biomedizinische Anwendungen

Fibrous proteins display different sequences and structures that have been used for various applications in biomedical fields such as biosensors, ...

Fabrication of a Bioactive, PCL-based "Self-fitting" Shape Memory Polymer Scaffold

Tissue engineering has been explored as an alternative strategy for the treatment of critical-sized cranio-maxillofacial (CMF) bone defects. Essential to the success of this approach is a scaffold that is able to conformally fit within an irregular defect while also having the requisite biodegradability, pore interconnectivity and bioactivity. By nature of their shape recovery and fixity properties, shape memory polymer (SMP) scaffolds could achieve defect &#8220;self-fitting.&#8221; In this way, following exposure to warm saline (~60 &#186;C), the SMP scaffold would become malleable, permitting it to be hand-pressed into an irregular defect. Subsequent cooling (~37 &#186;C) would return the scaffold to its relatively rigid state within the defect. To meet these requirements, this protocol describes the preparation of SMP scaffolds prepared via the photochemical cure of biodegradable polycaprolactone diacrylate (PCL-DA) using a solvent-casting particulate-leaching (SCPL) method. A fused salt template is utilized to achieve pore interconnectivity. To realize bioactivity, a polydopamine coating is applied to the surface of the scaffold pore walls. Characterization of self-fitting and shape memory behaviors, pore interconnectivity and in vitro bioactivity are also described.

Fabrication of a Bioactive, PCL-based &quot;Self-fitting&quot; Shape Memory Polymer Scaffold

Scaffolds capable of fitting within cranio-maxillofacial (CMF) bone defects while exhibiting osteoconductivity and bioactivity are of interest. This protocol describes the preparation of a shape memory scaffold based on polycaprolactone diacrylate (PCL-DA) using a solvent-casting particulate-leaching (SCPL) method employing a fused salt template and application of a bioactive polydopamine coating.

Herstellung eines bioaktiven, PCL-basierte &quot;Self-Montage&quot; Formgedächtnispolymer Scaffold

Tissue engineering has been explored as an alternative strategy for the treatment of critical-sized cranio-maxillofacial (CMF) bone defects. Essential to ...

Fabricating Superhydrophobic Polymeric Materials for Biomedical Applications

Superhydrophobic materials, with surfaces possessing permanent or metastable non-wetted states, are of interest for a number of biomedical and industrial applications. Here we describe how electrospinning or electrospraying a polymer mixture containing a biodegradable, biocompatible aliphatic polyester (e.g., polycaprolactone and poly(lactide-co-glycolide)), as the major component, doped with a hydrophobic copolymer composed of the polyester and a stearate-modified poly(glycerol carbonate) affords a superhydrophobic biomaterial. The fabrication techniques of electrospinning or electrospraying provide the enhanced surface roughness and porosity on and within the fibers or the particles, respectively. The use of a low surface energy copolymer dopant that blends with the polyester and can be stably electrospun or electrosprayed affords these superhydrophobic materials. Important parameters such as fiber size, copolymer dopant composition and/or concentration, and their effects on wettability are discussed. This combination of polymer chemistry and process engineering affords a versatile approach to develop application-specific materials using scalable techniques, which are likely generalizable to a wider class of polymers for a variety of applications.

Two- and three-dimensional superhydrophobic polymeric materials are prepared by electrospinning or electrospraying biodegradable polymers blended with a lower surface energy polymer of similar composition.

Herstellung Superhydrophobe Polymere Materialien für biomedizinische Anwendungen

Superhydrophobic materials, with surfaces possessing permanent or metastable non-wetted states, are of interest for a number of biomedical and industrial ...

Fabrication of a Dipole-assisted Solid Phase Extraction Microchip for Trace Metal Analysis in Water Samples

This paper describes a fabrication protocol for a dipole-assisted solid phase extraction (SPE) microchip available for trace metal analysis in water samples. A brief overview of the evolution of chip-based SPE techniques is provided. This is followed by an introduction to specific polymeric materials and their role in SPE. To develop an innovative dipole-assisted SPE technique, a chlorine (Cl)-containing SPE functionality was implanted into a poly(methyl methacrylate) (PMMA) microchip. Herein, diverse analytical techniques including contact angle analysis, Raman spectroscopic analysis, and laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) analysis were employed to validate the utility of the implantation protocol of the C-Cl moieties on the PMMA. The analytical results of the X-ray absorption near-edge structure (XANES) analysis also demonstrated the feasibility of the Cl-containing PMMA used as an extraction medium by virtue of the dipole-ion interactions between the highly electronegative C-Cl moieties and the positively charged metal ions.

The fabrication protocol of a dipole-assisted solid phase extraction microchip for the trace metal analysis is presented.

Die Herstellung einer Dipole-unterstützte Festphasenextraktion Microchip für die Spurenanalyse in Wasserproben

This paper describes a fabrication protocol for a dipole-assisted solid phase extraction (SPE) microchip available for trace metal analysis in water ...

Rapid Mix Preparation of Bioinspired Nanoscale Hydroxyapatite for Biomedical Applications

Hydroxyapatite (HA) has been widely used as a medical ceramic due to its good biocompatibility and osteoconductivity. Recently there has been interest regarding the use of bioinspired nanoscale hydroxyapatite (nHA). However, biological apatite is known to be calcium-deficient and carbonate-substituted with a nanoscale platelet-like morphology. Bioinspired nHA has the potential to stimulate optimal bone tissue regeneration due to its similarity to bone and tooth enamel mineral. Many of the methods currently used to fabricate nHA both in the laboratory and commercially, involve lengthy processes and complex equipment. Therefore, the aim of this study was to develop a rapid and reliable method to prepare high quality bioinspired nHA. The rapid mixing method developed was based upon an acid-base reaction involving calcium hydroxide and phosphoric acid. Briefly, a phosphoric acid solution was poured into a calcium hydroxide solution followed by stirring, washing and drying stages. Part of the batch was sintered at 1,000 &#176;C for 2 h in order to investigate the products&#39; high temperature stability. X-ray diffraction analysis showed the successful formation of HA, which showed thermal decomposition to &#946;-tricalcium phosphate after high temperature processing, which is typical for calcium-deficient HA. Fourier transform infrared spectroscopy showed the presence of carbonate groups in the precipitated product. The nHA particles had a low aspect ratio with approximate dimensions of 50 x 30 nm, close to the dimensions of biological apatite. The material was also calcium deficient with a Ca:P molar ratio of 1.63, which like biological apatite is lower than the stoichiometric HA ratio of 1.67. This new method is therefore a reliable and far more convenient process for the manufacture of bioinspired nHA, overcoming the need for lengthy titrations and complex equipment. The resulting bioinspired HA product is suitable for use in a wide variety of medical and consumer health applications.

This paper describes a novel method for the rapid manufacture of high quality bioinspired nanoscale hydroxyapatite. This biomaterial is of great significance in the manufacture of a wide range of innovative medical devices for clinical applications in orthopedics, craniofacial surgery and dentistry.

Rapid-Mix Herstellung von Bioinspirierte Nanoskalige Hydroxyapatit für biomedizinische Anwendungen

Hydroxyapatite (HA) has been widely used as a medical ceramic due to its good biocompatibility and osteoconductivity. Recently there has been interest ...

Patterning Bioactive Proteins or Peptides on Hydrogel Using Photochemistry for Biological Applications

There are many biological stimuli that can influence cell behavior and stem cell differentiation. General cell culture approaches rely on soluble factors within the medium to control cell behavior. However, soluble additions cannot mimic certain signaling motifs, such as matrix-bound growth factors, cell-cell signaling, and spatial biochemical cues, which are common influences on cells. Furthermore, biophysical properties of the matrix, such as substrate stiffness, play important roles in cell fate, which is not easily manipulated using conventional cell culturing practices. In this method, we describe a straightforward protocol to provide patterned bioactive proteins on synthetic polyethylene glycol (PEG) hydrogels using photochemistry. This platform allows for the independent control of substrate stiffness and spatial biochemical cues. These hydrogels can achieve a large range of physiologically relevant stiffness values. Additionally, the surfaces of these hydrogels can be photopatterned with bioactive peptides or proteins via thiol-ene click chemistry reactions. These methods have been optimized to retain protein function after surface immobilization. This is a versatile protocol that can be applied to any protein or peptide of interest to create a variety of patterns. Finally, cells seeded onto the surfaces of these bioactive hydrogels can be monitored over time as they respond to spatially specific signals.

In this method, we use photopolymerization and click chemistry techniques to create protein or peptide patterns on the surface of polyethylene glycol (PEG) hydrogels, providing immobilized bioactive signals for the study of cellular responses in vitro.

Musterung bioaktive Proteine oder Peptide auf Hydrogel mit Photochemie für biologische Anwendungen

There are many biological stimuli that can influence cell behavior and stem cell differentiation. General cell culture approaches rely on soluble factors ...

Melt Electrospinning Writing of Three-dimensional Poly(&#949;-caprolactone) Scaffolds with Controllable Morphologies for Tissue Engineering Applications

This tutorial reflects on the fundamental principles and guidelines for electrospinning writing with polymer melts, an additive manufacturing technology with great potential for biomedical applications. The technique facilitates the direct deposition of biocompatible polymer fibers to fabricate well-ordered scaffolds in the sub-micron to micro scale range. The establishment of a stable, viscoelastic, polymer jet between a spinneret and a collector is achieved using an applied voltage and can be direct-written. A significant benefit of a typical porous scaffold is a high surface-to-volume ratio which provides increased effective adhesion sites for cell attachment and growth. Controlling the printing process by fine-tuning the system parameters enables high reproducibility in the quality of the printed scaffolds. It also provides a flexible manufacturing platform for users to tailor the morphological structures of the scaffolds to their specific requirements. For this purpose, we present a protocol to obtain different fiber diameters using melt electrospinning writing (MEW) with a guided amendment of the parameters, including flow rate, voltage and collection speed. Furthermore, we demonstrate how to optimize the jet, discuss often experienced technical challenges, explain troubleshooting techniques and showcase a wide range of printable scaffold architectures.

Melt Electrospinning Writing of Three-dimensional Poly(&amp;#949;-caprolactone) Scaffolds with Controllable Morphologies for Tissue Engineering Applications

This protocol serves as a comprehensive guideline to fabricate scaffolds via electrospinning with polymer melts in a direct writing mode. We systematically outline the process and define the appropriate parameter settings for achieving targeted scaffold architectures.

Elektrospinnen Schreiben von dreidimensionalen Poly(ε-caprolactone) Gerüste mit steuerbaren Morphologien für Tissue Engineering Anwendungen zu schmelzen

This tutorial reflects on the fundamental principles and guidelines for electrospinning writing with polymer melts, an additive manufacturing technology ...

Fabrication and Characterization of Griffithsin-modified Fiber Scaffolds for Prevention of Sexually Transmitted Infections

Electrospun fibers (EFs) have been widely used in a variety of therapeutic applications; however, they have only recently been applied as a technology to prevent and treat sexually transmitted infections (STIs). Moreover, many EF technologies focus on encapsulating the active agent, relative to utilizing the surface to impart biofunctionality. Here we describe a method to fabricate and surface-modify poly(lactic-co-glycolic) acid (PLGA) electrospun fibers, with the potent antiviral lectin Griffithsin (GRFT). PLGA is an FDA-approved polymer that has been widely used in drug delivery due to its outstanding chemical and biocompatible properties. GRFT is a natural, potent, and safe lectin that possesses broad activity against numerous viruses including human immunodeficiency virus type 1 (HIV-1). When combined, GRFT-modified fibers have demonstrated potent inactivation of HIV-1 in vitro. This manuscript describes the methods to fabricate and characterize GRFT-modified EFs. First, PLGA is electrospun to create a fiber scaffold. Fibers are subsequently surface-modified with GRFT using 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS)chemistry. Scanning electron microscopy (SEM) was used to assess the size and morphology of surface-modified formulations. Additionally, a gp120 or hemagglutinin (HA)-based ELISA may be used to quantify the amount of GRFT conjugated to, as well as GRFT desorption from the fiber surface. This protocol can be more widely applied to fabricate fibers that are surface-modified with a variety of different proteins.

This manuscript describes the procedure to fabricate and characterize Griffithsin-modified poly(lactic-co-glycolic acid) electrospun fibers that demonstrate potent adhesive and antiviral activity against human immunodeficiency virus type 1 infection in vitro. Methods used to synthesize, surface-modify, and characterize the resulting morphology, conjugation, and desorption of Griffithsin from surface-modified fibers are described.

Herstellung und Charakterisierung von Griffithsin-modifizierte Faser Gerüste für die Prävention von sexuell übertragbare Infektionen

Electrospun fibers (EFs) have been widely used in a variety of therapeutic applications; however, they have only recently been applied as a technology to ...