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).

Gözenekli Metal iskelelerinin 1. Fabrikasyon <ol><li> Dört başlangıç ​​gözenek (40, 50, 60 ve 70), gözenekli Ti iskeleleri için Tablo 1 de tarif edildiği gibi malzemeler, uygun miktarlarda tartı sonra ticari olarak temin edilebilir Ti tozu, kamfen ve KD-4 karıştırılarak Ti-kamfen bulamaçlar hazırlayın. 500 ml polietilen (PE) şişelere çamurlar dökün ve 30 rpm&#39;de bir bilyeli değirmen fırın içinde 30 dakika boyunca 55 ° C&#39;de şişeler döndürün. </li><li> Silindirik alüminyum (Al) 60 mm&#39;lik bir çapa ve 60 mm yüksekliğe sahip kalıplar içine PE şişe çamurlar dökün. Karşılık gelen Al kayar kapak ile birlikte, Al kalıp kapatın ve 10 dakika süreyle 55 ° C &#39;de 30 rpm bir hızda bir bilyalı değirmen fırında kalıpları döndürün. <ol><li> Daha sonra, 44 ° C&#39;ye kadar bilyalı değirmen fırının sıcaklığı azaltmak ve sürekli 12 saat boyunca 44 ° C sabit sıcaklıkta 30 rpm bir hızda kalıp döndürün. </li></ol></li><li> Ball- kalıp dışarı atınDeğirmen fırın sonra ilave bir soğutma işlemi için 1 saat süre ile oda sıcaklığında kalıp döner. Bir Al dalgıç kullanarak Al kalıp katılaşmış titanyum / kamfen yeşil gövdesini sökün. </li><li> Elle bir lastik torba içinde katılaşmış titanyum / kamfen yeşil vücudu yerleştirin ve tamamen bir dize ile torbanın ağzını bağlayarak lastik torbanın ağzını sıkıca kapatın. Soğuk izostatik presleme (CIP) makinenin su tankı kauçuk torba yerleştirin ve 10 dakika için 200 MPa, eşbasınçlı bir basınç uygulanır. Lastik torbadan sıkıştırılmış yeşil vücudu çıkarın. </li><li> Elle bir alümina pota üzerine Ti-kamfen yeşil vücudu aktarın ve dondurarak kurutma makinesinde pota yerleştirin. Dondurularak kuru yeşil vücutta kamfen fazı süblimleşme yeşil gövde - 24 saat boyunca 40 ° C. </li><li> Daha sonra, bir alümina kapak kayma Kroze kapatın ve oda sıcaklığında (10 -6 Torr altında) bir vakum fırınında kapalı pota yerleştirin. Sonra, bir ısıtma r 1300 ° C fırının sıcaklığını artırmak5 ° C / dk arasında yedi ve 2 saat süre ile 1300 ° C sıcaklık tutun. </li><li> Isıl işlemden sonra, fırının tam oda sıcaklığına kadar soğutuldu kadar 6-7 saat boyunca fırında sinterlenmiş gözenekli Ti tutun. Not: Soğutma işlemi 6 saat süresince, 400 ° C&#39;nin üzerinde fırının ortalama soğutma hızı ~ 15 ° C / dk ve 400 ° C&#39;nin altında fırının ortalama soğutma hızı ~ 2 ° C / dk. </li><li> Gerekirse, elektriksel deşarj işlemesi ile 16 mm (EDM) bir çapa sahip olan bir disk şeklinde numuneler halinde sinterlenmiş gözenekli Ti blok kesin. 27 Not: Al kalıp boyutuna bağlı olarak tabaka, gözenekli Ti boyutu işleme süreci (Şekil 2A) ile değiştirilmesi gerekir. </li><li> Bir otoklav içinde, gözenekli Ti örnekleri bir cam behere koyun ve 15 dakika boyunca 121 ° C&#39;de sterilize örnekleri. Numuneler otoklavdan çıkarılmaktadır. Iki kere% 70 etanol ile iki kez ve sonra damıtılmış su ile gözenekli Ti örnekleri yıkayın.Son olarak, Petri kabı içine gözenekli Ti bırakın ve UV ışığı altında temiz bir bankta oda sıcaklığında örnekleri Hava kurulayın. </li></ol> Biyoaktif Ajanların ile iskelelerinin 2. Dip Kaplama <ol><li> 9 Dulbecco Fosfat Tamponlu Salin (DPBS, pH 7.4) bir solüsyonu 10 ml bir ml sterilize olan GFP 1 ml karıştırılarak temiz bir tezgah 100 ug / ml 1 mg / ml, ticari yeşil floresans proteini (GFP) seyreltin Tablo 1 &#39;de gösterildiği gibi, polistiren (PS), tüpü. </li><li> Oda sıcaklığında GFP çözeltisi ile PS tüpüne Ti numuneler yerleştirilerek ve temiz bir tezgah üzerine yerleştirerek seyreltildi GFP çözeltisi (100 ug / ml), 10 ml sterilize edilmiş, yoğun ya da gözenekli Ti bırakın. </li><li> Bir vakum kurutucu içinde PS tüp yerleştirin ve 10 dakika GFP çözeltisi daha etkili bir şekilde, gözenekli Ti gözeneklerinden içeri girmesi ile sağlamak için desikatörde tahliye. </li><li> Cımbız kullanarak PS tüpünden gözenekli titanyum çıkarın. 10 cm çapında Pe içine GFP kaplamalı gözenekli Ti yerleştirinÜç çanak ve hava-kuru O / N temiz bir bankta RT de. </li><li> Bir cam beher içinde Dulbecco Fosfat Tamponlu Salin (DPBS) 10 ml iki kez gözenekli Ti durulayın ve temiz bir tezgah üzerinde RT cımbız ve hava kuru kullanılarak 10 cm çapında bir Petri kabı içine, gözenekli Ti hareket eder. </li></ol> Gözenekli iskelelerinin 3. Yoğunlaştırma <ol><li> Silindirik çelik kalıpta çeşitli yükseklikleri GFP kaplı gözenekli Ti örnekleri yerleştirin ve çelik kalıbın (Şekil 3A) üst ve alt deliklere yumruklar bir dizi yerleştirin. </li><li> 0.05 ~ 0.1 sn ara gerilme oranlarında bir basın makine kullanarak örnek (Şekil 3A) z yönünde oda sıcaklığında çelik kalıp montaj içinde gözenekli Ti sıkıştır -1 Tablo 2&#39;de gösterilen önceden belirlenmiş uygulanan suşlarına karşı. Basınç tutun boşaltma öncesinde 1 dakika karıştırıldı. </li><li> Çelik kalıbından yoğunlaştırılmış Ti örnekleri çıkarın. DPBS 10 ml iki kez yoğunlaştırılmış numune yıkayınTemiz bir bankta oda sıcaklığında bir beher ve hava kuru O / N. </li></ol> GFP-kaplı iskelelerinin 4. Salınım test <ol><li> Numunelerin üç tip daldırın (GFP-kaplanmış, yoğun Ti (adımlardan sonra 2), GFP-kaplanmış gözenekli Ti (Adım 1 ve 2) ve GFP-kaplı yoğunlaştırılmış gözenekli Ti (adım 1-3) sonrası) sonra 5 ml DPBS (pH Temiz bir tezgah üzerinde 37 ° C&#39;de 10 mi steril PS tüp içinde ihtiva 7,4) çözeltisi eklendi. </li><li> Ve GFP-kaplanmış numune, her PS borusundan Emme bütün DPBS çözeltisi, 1 önceden belirlenen bir, 2, 3, 5, 8, 12 e göre bir pipet kullanarak yeni bir 5 ml DPBS çözeltisi (pH 7,4) ile yeniden doldurmak batırıldıktan sonra 15, 22 ve 29 gün. </li><li> Daldırma (gün 0) önce, GFP-kaplanmış numunelerin parlaklığı görüntü almak ve 22 gün batırıldıktan sonra konfokal lazer tarama spektroskopisi (CLSM) ile. </li><li> Kullanılarak bölüm 4.2&#39;de her PS tüpten çekilir 5 mi DPBS çözeltisinin bir toplam 1 ml çözelti içinde serbest GFP floresans sinyali yoğunluğu ölçmek215 nm dalga boyunda UV spektroskopisi. Standart eğri kullanılarak GFP çözeltisinin konsantrasyonu yoğunluk değeri dönüştürün. Not: Ölçme önce, 0 ng / ml konsantrasyon aralığında, GFP çözeltisi floresans sinyali yoğunluğu ölçülerek GFP çözeltisi standart eğri çizmek - 10 ug / ml olmuştur. </li></ol> Kademeli Gözenekli Ti iskelelerinin 5. Fabrikasyon <ol><li> 1,7 adıma adım 1.1 tekrarlayarak sinterlenmiş gözenekli Ti bloğunu üretin. </li><li> Makine EDM tarafından önceden belirlenmiş bir yapı tasarımları (örneğin, Şekil 5a ve 5d) göre sinterlenmiş gözenekli Ti bloğu. </li><li> Gözenekli Ti çapıdır ~ kalıbın çapından 0,1 mm küçük çelik bir kalıp yükseklik dağılımı ile işlenmiş Ti numuneler ve çelik kalıbın üst ve alt deliklere zımbaların kümesini yerleştirebilir. </li><li> Adımları 3.2 ve 3.3 gerçekleştirin. </li></ol> 6. Porozite MeTi iskelelerinin asurement <ol><li> Ti iskelelerinin kitle (m s) ölçün. </li><li> Ti iskelelerinin uzunluk, genişlik ve yükseklik ölçerek belirgin hacmi (V s) hesaplayın. </li><li> Aşağıdaki denklem kullanılarak gözeneklilik hesaplamak: <img alt="Denklem 1" src="/files/ftp_upload/53279/53279eq1.jpg" /> P toplam porozite yüzdesi, ρ Ti titanyum ve m S teorik yoğunluğu olduğu / V S numunenin ölçülen yoğunluğudur. Not: MikroBT&#39;lerin görüntüleme, bir mikro-bilgisayarlı tomografi kullanılarak gerçekleştirilir sonra Ti örnekleri gözenekliliği ile doğrudan MikroBT&#39;lerin görüntüleri elde edilebilir. </li></ol>

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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porous+titanium+(Ti)+scaffolds+by+freezing+TiH2/camphene+slurries.">Porous titanium (Ti) scaffolds by freezing TiH2/camphene slurries.</a> Mater. Lett. 62 (30), 4506-4508 (2008).</li><li>Yook, S. W., Kim, H. E., Koh, Y. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fabrication+of+porous+titanium+scaffolds+with+high+compressive+strength+using+camphene-based+freeze+casting.">Fabrication of porous titanium scaffolds with high compressive strength using camphene-based freeze casting.</a> Mater. Lett. 63 (17), 1502-1504 (2009).</li></ol>

The authors declare that they have no competing financial interests.

BIOMETAL sistemleri yaygın biyomedikal uygulamalar için kullanılmış olsa da, özellikle de, taşıyıcı malzemeler olarak, yüksek sertlik ve metal düşük biyo gibi büyük zorluklar olarak kabul edilmiştir. Bu çalışmada, yeni bir metal sistemi, biyomimetik mekanik özellikleri yanı sıra sürdürülebilir bırakma davranışı ile biyoaktif yüzeye sahip bir yoğunlaştırılmış gözenekli metal iskele imalat yöntemi kurdu. Bizim fabrikasyon yönteminin önemli avantajları 1) biz zaten geliştirilen ö...

Metalik biyomalzemeler yaygın nedeniyle mükemmel mekanik mukavemet ve esneklik, 1-3 yük taşıyan implantlar ve internal fiksasyon cihazları olarak kullanılmış olsa da iki kritik zorlukları içerir: 1) Mekanik uyuşmazlığı metaller biyolojik dokulara daha sert olduğu için, istenmeyen hasara yol genellikle yabancı cisim reaksiyonları (örneğin, enflamasyon veya tromboz) provoke çevre dokulara ve genellikle biyolojik dokularla yoksul bir arayüz ile sonuçlanır 2) düşük biyo-için. 4-6 Gözenekli metal iskeleleri iyileştirilmesi, yapılarda kemik büyümesini teşvik etmek için ileri sürülmüştür . Stres kalkan etkileri nedeniyle azaltılmış sertlik bastırılır ise kemik-implant temas 7-9 Dahası, çeşitli yüzey modifikasyonlar metalik implantların biyolojik aktivitelerini geliştirmek için uygulanmıştır; Bu tür modifikasyonlar, kaplamaya biyoaktif moleküllerin (örneğin, büyüme faktörleri için metal yüzeye sahiptirları) ya da ilaçlar (örn, vankomisin, tetrasiklin). 10-12 Ancak, bu tür gözenekli metal iskelelerinin azaltılmış mekanik özellikleri gibi sorunlar, sertlik ve biyoaktif kaplama tabakalarının hızlı salım hâlâ çözülememiştir azalmıştır. 13-16 Özellikle, titanyum (Ti) ve Ti alaşımları için, mükemmel mekanik özellikleri, kimyasal stabilite en popüler BIOMETAL sistemlerinden biri ve iyi biyolojik uygunluktur bulunmaktadır. 13,17-19 Bunların köpük şeklindeki uygulamalar da çeken 3D için artan bir ilgi gözenekli ağlar 20-22 çabalar polimerik sünger çoğaltma, metal parçacıklar, hızlı prototipleme (RP) yönteminin sinterleme dahil olmak üzere yeni üretim tekniklerini geliştirerek mekanik özelliklerini geliştirmek için yapılmıştır. kemik gibi mekanik özelliklerinin yanı sıra, kemik büyümesini teşvik ve gözenekler çeşitli özelliklerini kontrol etmek için uzay tutucu yöntemi (örneğin, gözenek fraksiyonu,şekil, boyut, dağıtım ve bağlantı) ve malzeme özellikleri (örneğin, metalik faz ve kirlilik). Son zamanlarda, su bazlı metal çamurunun donma döküm iyi hizalanmış gözenek ile mekanik gelişmiş Ti formlarını üretmek için büyük ilgi görmüştür 23-25 katılaşma sırasında tek yönlü buz dendrit büyümesini kullanarak yapılar; Ancak, su ile metal tozlarının temasından kaynaklanan oksijen kirlenme Ti iskelelerinin kırılganlığını en aza indirmek için özel bir bakım gerektirir. 14,15 Bu nedenle, biyolojik olarak aktif ve mekanik ayarlanabilir gözenekli Ti iskeleleri imal yönelik yeni bir yaklaşım geliştirmişlerdir. 25 iskeleleri, başlangıçta% 50&#39;nin üzerinde bir gözenekliliğe sahip, gözenekli bir yapıya sahiptir. Imal gözenekli iskeleleri sonuçta elde edilen gözeneklilik, mekanik özellikler ve ilaç salım davranışı başvurunun ile kontrol edildi sırasında mekanik bir pres kullanılarak biyolojik olarak aktif moleküller ile kaplanır ve daha sonra sıkıştırılaned süzün. Yoğunlaştırılmış gözenekli Ti implantlar kemik (3-20 GPa). 2 için kaplama tabakasının karşılaştırılabilir düşük sertliğe rağmen, iyi bir mukavemete sahip düşük bir gözenekliliğe göstermiştir, yoğunlaştırılmış gözenekli Ti biyo-etkinliği önemli ölçüde geliştirilmiştir. Üstelik, yoğunlaştırma işlemi ile oluşturulan eşsiz düz gözenek yapıları kaplanmış biyoaktif molekülleri yavaş yavaş, uzun bir süre boyunca etkinliğini muhafaza iskele açıklanacak görülmüştür. Bu çalışmada, biyomedikal uygulamalarda potansiyel kullanım için yoğunlaştırılmış gözenekli Ti iskeleleri imal etmek bizim kurulan yöntemi tanıttı. Protokol metal çamurları ve gözenekli iskeleleri sıklaştırılması ile dinamik donma döküm içerir. İlk olarak, Şekil 1A gösterildiği gibi dinamik donma döküm yöntemi tanıtıldı iyi süneklik ile gözenekli Ti iskeleleri imal etmek. Ti tozu, sıvı kamfen içinde dağıtılır; daha sonra, sıcaklık düşürülerekSıvı faz Ti tozu ağı ve bir katı kamfen kristalleri arasında faz ayrımı ile sonuçlanan, katılaştırıldı. Daha sonra, katılaşmış Ti-kamfen yeşil gövde içinde Ti tozlar, sürekli Ti dikmeler ile kondanse edildi sinterlenmiştir ve kamfen fazı tamamen gözenekli bir yapı elde etmek için uzaklaştırılmıştır. Elde edilen gözenekli iskeleleri ile kaplama ve yoğunlaştırma işlemi sıklaştırılması ve ilk gözeneklilik derecesi değişen kullanılmıştır. Kaplama tabakası ve serbest bırakma davranışı görüntülendi ve yeşil flüoresan proteini (GFP) GFP-kaplanmış, yoğun Ti göre ve yoğunlaştırma, gözenekli Ti -kaplı kullanılarak ölçüldü. Son olarak, iki farklı gözenekli yapılara sahiptir fonksiyonel aşamalı Ti iskeleleri önerilen ve gözenekli iskeleler iç ve dış parçaları yoğunlaştırılması derecesini değiştirmek suretiyle gösterildi.

<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.

Gözenekli Ti iskeleleri üretilmesi için kullanılan imalat süreci Şekil 1A&#39;da gösterilmiştir. Ti tozu, sıvı kamfen tamamen katılaştıktan ise, nispeten ağır Ti tozunun herhangi bir çökeltiler minimize edilebilir, 12 saat boyunca 44 ° C&#39;de kabın sürekli dönüşüyle ​​kamfen içinde homojen biçimde dağılmış ve tutulur. Burada 3-boyutlu birbirine büyük kamfen gözenekler Ti tozu faz (Şekil 1C) ile çevrilidir, Şekil 1B</stro...

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.

Biyomedikal Uygulamalar için Mekanik ayarlanabilir ve Biyoaktif Metal iskelelerinin Fabrikasyon

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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10.6K Görüntüleme.  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.

Mekanik Dinamik Hücre Kültürü için Microfabricated platformlar

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

Biyomühendislik

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.

3D Fibrin hazırlanması Kök Hücre Kültürü Uygulamaları için iskeleleri

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.

Biyomedikal Uygulamalar için İpek-ipek Protein Alaşım Materyalleri Tasarlama

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.

Bir Biyoaktif, PCL-temelli &quot;Kendinden uydurma&quot; Shape Memory Polymer İskele imalatı

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.

Biyomedikal Uygulamalar için Süperhidrofobik Polimer Malzemeler imalatı

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.

Su Örneklerinde Eser Metal Analizi için Dipol destekli Katı Faz Ekstraksiyon Microchip Fabrikasyon

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.

Biyomedikal Uygulamalar için Biyolojik Tabanlı Akıllı Nano Hidroksiapatit Rapid Mix Hazırlanması

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.

Biyoaktif proteinler veya peptidler hidrojel Photochemistry biyolojik uygulamalarda kullanarak üzerinde desenlendirme

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.

Üç boyutlu Poly(ε-caprolactone) İskele kontrol edilebilir türleri morfoloji ile Electrospinning yazma doku mühendisliği uygulamaları için erime

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.

İmalat ve Fiber Griffithsin-modified iskele karakterizasyonu önlenmesi için cinsel enfeksiyonlar iletilen

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