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. Realizzazione di metallo poroso Ponteggi <ol><li> Preparare fanghi Ti-canfene mescolando disponibile in commercio in polvere Ti, canfene, e KD-4 dopo aver valutato l&#39;ammontare adeguato di materiali come descritto nella tabella 1 per porosi ponteggi TI con quattro porosità iniziale (40, 50, 60, e 70). Versare fanghi in 500 ml di polietilene (PE) e ruotare le bottiglie bottiglie a 55 ° C per 30 minuti in un forno di macinazione a 30 rpm. </li><li> Versare fanghi dalle bottiglie PE in alluminio cilindrico (Al) stampi con un diametro di 60 mm ed un&#39;altezza di 60 mm. Sigillare ogni stampo Al con la corrispondente copertura di slittamento Al e ruotare gli stampi in forno di macinazione ad una velocità di 30 giri al minuto a 55 ° C per 10 min. <ol><li> Successivamente, diminuire la temperatura del forno di macinazione a 44 ° C, e continuamente ruotare gli stampi ad una velocità di 30 giri al minuto a temperatura costante di 44 ° C per 12 ore. </li></ol></li><li> Togliere lo stampo dal palla-mulino forno dopo inoltre ruotare gli stampi a temperatura ambiente per 1 ora per il processo di raffreddamento. Rimuovere il titanio / canfene corpo verde solidificato dallo stampo di Al con un pistone di Al. </li><li> Posizionare il titanio / canfene corpo verde solidificato in un sacchetto di gomma a mano e completamente sigillare il sacchetto di gomma legando la bocca del sacco con una stringa. Posizionare il sacchetto di gomma nel serbatoio di una pressatura isostatica (CIP) Macchina fredda e applicare una pressione isostatica di 200 MPa per 10 min. Rimuovere il corpo verde compressa dal sacchetto di gomma. </li><li> Trasferire il corpo verde Ti-canfene su un crogiolo di allumina a mano e posizionare il crogiolo nella macchina liofilizzatore. Liofilizzare il corpo verde sublimare fase canfene nel corpo verde a - 40 ° C per 24 ore. </li><li> Successivamente, chiudere il crogiolo con una copertura di slittamento allumina e posizionare il crogiolo chiuso in un forno sotto vuoto (inferiore a 10 -6 Torr) a temperatura ambiente. Quindi, aumentare la temperatura del forno a 1300 ° C ad un riscaldamento rate di 5 ° C / min e mantenere la temperatura a 1300 ° C per 2 ore. </li><li> Dopo il trattamento termico, mantenere la sinterizzato poroso Ti nel forno per 6-7 ore finché il forno è completamente raffreddata a RT. Nota: Durante 6 hr del processo di raffreddamento, la velocità di raffreddamento media del forno sopra 400 ° C è di ~ 15 ° C / min e la velocità di raffreddamento media del forno inferiore a 400 ° C è ~ 2 ° C / min. </li><li> Se necessario, tagliare il blocco di sinterizzato poroso Ti in campioni a forma di disco con un diametro di 16 mm mediante elettroerosione (EDM). 27 Nota: A seconda delle dimensioni degli stampi Al, la dimensione del sinterizzato poroso Ti deve essere modificata durante il processo di lavorazione (Figura 2A). </li><li> Posizionare un bicchiere di vetro con i campioni porosi Ti in autoclave e sterilizzare i campioni a 121 ° C per 15 min. Rimuovere i campioni dall&#39;autoclave. Lavare i campioni porosi TI con acqua distillata e quindi due volte con etanolo al 70% per due volte.Infine, lasciare la porosa Ti in una capsula di Petri e aria asciugare i campioni a temperatura ambiente su una panchina pulita sotto raggi UV. </li></ol> 2. Immergere Rivestimento di impalcature con gli agenti bioattivi <ol><li> Diluire il commerciale verde Fluorescence Protein (GFP) da 1 mg / ml a 100 pg / ml in un banco pulito mescolando 1 ml di GFP con 9 ml di soluzione di Dulbecco Phosphate Buffered Saline (DPBS, pH 7.4) in 10 ml-sterilizzati polistirene (PS) tubo come indicato nella Tabella 1. </li><li> Immergere il sterilizzato densa o porosa Ti in 10 ml di soluzione diluita GFP (100 mcg / ml) ponendo i campioni di Ti nel tubo PS con la soluzione GFP a RT e mettendo su un banco pulito. </li><li> Posizionare il tubo PS in un essiccatore a vuoto ed evacuare l&#39;essiccatore per 10 minuti per assicurare la soluzione GFP penetra più efficacemente i pori del porosa Ti. </li><li> Rimuovere il titanio poroso dal tubo di PS con una pinzetta. Posizionare la GFP rivestite poroso Ti in un diametro di 10 centimetri Pepiatto tri e asciugare O / N a RT su un banco pulito. </li><li> Sciacquare la porosa Ti due volte con 10 ml di Dulbecco tampone fosfato (DPBS) in un bicchiere di vetro, e spostare il porosa Ti in un diametro di Petri piatto 10 centimetri con una pinzetta e aria secca a temperatura ambiente su una panchina pulita. </li></ol> 3. densificazione porosi Ponteggi <ol><li> Porre i campioni Ti porosi GFP rivestite con diverse altezze in uno stampo cilindrico di acciaio, e inserire una serie di punzoni nei fori superiori e inferiori dello stampo in acciaio (Figura 3A). </li><li> Comprimere il porosa Ti nell&#39;assieme stampo di acciaio a temperatura ambiente nella direzione z del campione (Figura 3A) utilizzando una pressa a velocità di deformazione intermedie di 0,05 ~ 0,1 sec -1 contro le sollecitazioni applicate predeterminati illustrati nella Tabella 2. Tenere la pressione per 1 min prima di scarico. </li><li> Rimuovere i campioni Ti densificati dallo stampo in acciaio. Lavare i campioni addensate due volte con 10 ml di DPBSin un becher e asciugare O / N a RT su un banco pulito. </li></ol> 4. Prova Rilascio di Ponteggi GFP rivestite <ol><li> Immergere tre tipi di campioni (GFP rivestite denso Ti (dopo i passaggi da 2), rivestito GFP-poroso Ti (dopo i punti 1 e 2) e GFP-rivestito addensato poroso Ti (dopo i passi 1-3)) in 5 ml DPBS (pH 7.4) soluzione contenuta in una provetta da 10 ml sterilizzato PS a 37 ° C su un banco pulito. </li><li> Aspirazione su tutta la soluzione DPBS da ciascuna provetta PS con il campione GFP rivestite e riempite con una soluzione nuova 5 ml DPBS (pH 7,4) usando una pipetta secondo i tempi prestabiliti di 1, 2, 3, 5, 8, 12, 15, 22 e 29 giorni dopo l&#39;immersione. </li><li> Prendete le immagini di fluorescenza dei campioni GFP rivestite prima immersione (giorno 0) e dopo 22 giorni-immersion utilizzando la spettroscopia confocale a scansione laser (CLSM). </li><li> Misurare l&#39;intensità del segnale di fluorescenza della GFP rilasciato in 1 ml di soluzione da un totale di 5 ml di soluzione DPBS disegnata da ciascuna provetta PS nella sezione 4.2 usandoSpettroscopia UV alla lunghezza d&#39;onda di 215 nm. Convertire il valore di intensità nella concentrazione della soluzione GFP utilizzando la curva standard. Nota: Prima della misurazione, tracciare la curva standard di soluzione GFP misurando l&#39;intensità segnale di fluorescenza della soluzione GFP in un intervallo di concentrazione da 0 ng / ml - 10 mg / ml. </li></ol> 5. Realizzazione di Graded porosi Ti Ponteggi <ol><li> Produrre un blocco del sinterizzato poroso Ti ripetendo il punto 1.1 al punto 1.7. </li><li> Macchina del blocco poroso sinterizzato Ti secondo i disegni struttura predeterminata (ad esempio, la figura 5a e 5d) di EDM. </li><li> Porre i campioni lavorati TI con distribuzione dell&#39;altezza in uno stampo in acciaio, dove il diametro porosa Ti è ~ 0,1 mm inferiore al diametro dello stampo e inserire una serie di punzoni nei fori superiori e inferiori dello stampo in acciaio. </li><li> Eseguire i passi 3.2 e 3.3. </li></ol> 6. porosità Measurement di Ti Ponteggi <ol><li> Misurare la massa (m s) di ponteggi Ti. </li><li> Calcolare il volume apparente (V s), misurando la lunghezza, la larghezza e l&#39;altezza di scaffold Ti. </li><li> Calcolare la porosità utilizzando la seguente equazione: <img alt="Equazione 1" src="/files/ftp_upload/53279/53279eq1.jpg" /> dove P è la percentuale totale di porosità, ρ Ti è la densità teorica del titanio e m S / V S è la densità misurata del campione. Nota: La porosità dei campioni di Ti può essere recuperato direttamente dalle immagini microCT dopo l&#39;imaging microCT viene effettuata utilizzando una tomografia scanner micro-calcolata. </li></ol>

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The authors declare that they have no competing financial interests.

Mentre i sistemi Biometal sono stati ampiamente utilizzati per applicazioni biomediche, in particolare, come materiali portanti, elevata rigidità e bassa bioattività dei metalli sono stati considerati come grandi sfide. In questo studio, abbiamo stabilito il metodo di fabbricazione di un nuovo sistema di metallo, una impalcatura metallo poroso addensato che ha biomimetici proprietà meccaniche e di superficie bioattiva con comportamento rilascio sostenibile. I principali vantaggi del nostro metodo di fabbricazione inc...

Mentre biomateriali metallici sono stati ampiamente utilizzati come protesi portanti e dispositivi di fissaggio interni a causa della loro ottima resistenza meccanica e la resistenza, 1-3 coinvolgono due sfide cruciali: 1) non corrispondente meccanico perché i metalli sono molto più rigido rispetto ai tessuti biologici, provocando danni indesiderati ai tessuti circostanti e 2) bassa bioattività che spesso si traduce in scarsa interfaccia con i tessuti biologici, spesso provocando reazioni da corpo estraneo (ad esempio, infiammazione o trombosi). 4-6 ponteggi metallici porosi sono state proposte per promuovere la crescita ossea nelle strutture, migliorando . contatto osso-impianto, mentre gli effetti dello scudo di stress vengono soppressi a causa della loro rigidità ridotta 7-9 Inoltre, varie modifiche di superficie sono stati applicati per migliorare le attività biologiche di impianti metallici; tali modifiche comprendono il rivestimento della superficie metallica con molecole bioattive (ad esempio, la crescita factori) o farmaci (ad esempio, vancomicina, tetracicline). 10-12 Tuttavia, problemi come la riduzione delle proprietà meccaniche dei ponteggi metallici porosi, diminuito la rigidità e il rilascio veloce degli strati di rivestimento bioattivi restano irrisolti. 13-16 In particolare, titanio (Ti) e leghe di Ti sono uno dei più diffusi sistemi Biometal causa delle loro eccellenti proprietà meccaniche, stabilità chimica e buona biocompatibilità. 13,17-19 loro applicazioni a forma di schiuma hanno attirato un crescente interesse perché il 3D reti porosi promuovere crescita ossea in aggiunta alle proprietà meccaniche simile all&#39;osso. 20-22 sforzi sono stati fatti per migliorare le proprietà meccaniche sviluppando nuove tecniche di produzione inclusi replica di spugna polimerica, sinterizzazione di particelle metalliche, prototipazione rapida metodo (RP), e Metodo titolare spazio per controllare le varie caratteristiche dei pori (ad esempio, la frazione dei pori,forma, dimensione, di distribuzione, e la connettività) e proprietà dei materiali (ad esempio, di fase metallici e impurità). 23-25 ​​Recentemente, il casting congelamento dei a base di acqua metallo liquami ha guadagnato una notevole attenzione per la produzione di forme avanzate meccanicamente TI con pori ben allineati strutture, utilizzando la unidirezionale ghiaccio crescita dei dendriti durante la solidificazione; tuttavia, la contaminazione di ossigeno causata dal contatto di polveri di metallo con l&#39;acqua richiede una particolare attenzione per ridurre al minimo la fragilità di ponteggi Ti. 14,15 Pertanto, abbiamo sviluppato un nuovo approccio verso la fabbricazione di impalcature Ti porosi bioattive e meccanicamente accordabili. 25 Le impalcature hanno inizialmente strutture porose con una porosità superiore al 50%. Gli scaffold porosi fabbricati sono stati rivestiti con molecole bioattive e poi compressi utilizzando una pressa meccanica in cui la porosità finale, proprietà meccaniche e comportamenti rilascio del farmaco erano controllati dal appliceppo ed. Gli impianti addensate Ti porosi hanno dimostrato bassa porosità con buona resistenza nonostante la bassa rigidità paragonabile a quella dell&#39;osso (3-20 GPa). 2 A causa dello strato di rivestimento, la bioattività del addensato porosa Ti è significativamente migliorata. Inoltre, a causa delle particolari strutture a pori piane indotte dal processo di densificazione, le molecole bioattive rivestiti sono stati osservati essere gradualmente rilasciato dal ponteggio, mantenendo la loro efficacia per un periodo prolungato. In questo studio, abbiamo introdotto il nostro metodo stabilito per fabbricare densificati scaffold Ti porosi per uso potenziale in applicazioni biomediche. Il protocollo include colata congelamento dinamico con fanghi metallo e densificazione di scaffold porosi. In primo luogo, per fabbricare scaffold porosi TI con buona duttilità è stato introdotto il metodo freeze fusione dinamica come mostrato nella Figura 1A. Polvere ti è stata dispersa in camphene liquido; quindi, diminuendo la temperatura,la fase liquida è solidificato, causando la separazione di fase tra la rete polvere Ti e cristalli canfene solidi. Successivamente, il corpo verde solidificato Ti-canfene stato sinterizzato in cui le polveri di Ti sono stati condensati con continue puntoni Ti, e la fase canfene stato completamente rimosso per ottenere una struttura porosa. Il rivestimento e processo densificazione con scaffold porosi ottenuti è stato impiegato, variando il grado di addensamento e porosità iniziale. Lo strato di rivestimento ed il suo comportamento rilascio sono stati visualizzati e quantificati tramite la proteina fluorescente verde (GFP) Rivestiti poroso Ti con e senza densificazione rispetto alla GFP rivestite denso Ti. Infine, impalcature Ti funzionalmente graduati che hanno due diverse strutture porose sono stati proposti e dimostrate variando il grado di addensamento delle parti interne ed esterne dei scaffold porosi.

<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.

Il processo di fabbricazione utilizzata per produrre scaffold porosi Ti è illustrato nella Figura 1A. Polvere di Ti è mantenuto disperso omogeneamente in canfene dalla rotazione continua del contenitore a 44 ° C per 12 ore e, mentre canfene liquido è completamente solidificato, eventuali sedimenti di relativamente pesante polvere Ti possono essere minimizzati. Come risultato, il corpo verde omogeneo Ti-canfene stato prodotto utilizzando il processo di congelamento colata dinamica co...

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.

Fabbricazione di meccanicamente Tunable e Bioactive metallo Ponteggi per applicazioni biomediche

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

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Unit 1

Transformations of Stress and Strain

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10.6K Views.  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.

Video: 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.

Piattaforme microfabbricazione per coltura cellulare meccanicamente dinamica

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

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Bioingegneria

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.

Preparazione del 3D fibrina ponteggi per le applicazioni delle cellule staminali Cultura

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.

Progettazione di materiali in lega di proteine ​​della seta-seta per le applicazioni biomediche

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.

Realizzazione di una sede a PCL Bioactive, &quot;Self-fitting&quot; Shape Memory Polymer Ponteggio

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.

Realizzazione superidrofobiche Materiali Polimerici per applicazioni biomediche

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.

Realizzazione di un dipolo-assistita estrazione in fase solida Microchip per Trace analisi dei metalli in campioni di acqua

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 Preparazione di bioispirati Nanoscale idrossiapatite per applicazioni biomediche

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.

Patterning bioattivi proteine o peptidi su idrogel utilizzando fotochimica per applicazioni biologiche

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.

Sciogliere elettrofilatura scrittura di scaffold tridimensionale Poly(ε-caprolactone) con morfologie controllabile per applicazioni di ingegneria del tessuto

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.

Fabbricazione e caratterizzazione di fibra Griffithsin-modificato ponteggi per la prevenzione delle infezioni sessualmente trasmesse

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