Name: custom engineered tissue culture molds from laser-etched masters
Uploaded: 2018-05-21T13:00:00.000+00:00
Duration: 8 min 56 s
Description: Watch this Scientific Journal Video about Custom Engineered Tissue Culture Molds from Laser-etched Masters at JoVE.com

저자는 NIH R00 HL115123 및 브라운 대학 공학에서 자금을 인정 합니다. 그들은 또한 교육에 대 한 크리스 황소와 브라운 디자인 워크샵에 게 감사는 고 레이저 커터와 지원.

<ol>
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M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+antifibrinolytic+agents+on+the+life+span+of+fibrin+sealant.">Effects of antifibrinolytic agents on the life span of fibrin sealant.</a> J. Surg. Res. 53, 402-407 (1992).</li><li>Roberts, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stromal+Cells+in+Dense+Collagen+Promote+Cardiomyocyte+and+Microvascular+Patterning+in+Engineered+Human+Heart+Tissue.">Stromal Cells in Dense Collagen Promote Cardiomyocyte and Microvascular Patterning in Engineered Human Heart Tissue.</a> Tissue Eng. Part A. 22, 633-644 (2016).</li><li>Ye, K. Y., Sullivan, K. E., Black, L. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Encapsulation+of+Cardiomyocytes+in+a+Fibrin+Hydrogel+for+Cardiac+Tissue+Engineering.">Encapsulation of Cardiomyocytes in a Fibrin Hydrogel for Cardiac Tissue Engineering.</a> JoVE. , (2011).</li><li>Zimmermann, W. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue+Engineering+of+a+Differentiated+Cardiac+Muscle+Construct.">Tissue Engineering of a Differentiated Cardiac Muscle Construct.</a> Circ. Res. 90, 223-230 (2002).</li><li>McCain, M. L., Agarwal, A., Nesmith, H. W., Nesmith, A. P., Parker, K. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Micromolded+Gelatin+Hydrogels+for+Extended+Culture+of+Engineered+Cardiac+Tissues.">Micromolded Gelatin Hydrogels for Extended Culture of Engineered Cardiac Tissues.</a> Biomaterials. 35, 5462-5471 (2014).</li><li>Hu, J. J., Chen, G. W., Liu, Y. C., Hsu, S. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+Specimen+Geometry+on+the+Estimation+of+the+Planar+Biaxial+Mechanical+Properties+of+Cruciform+Specimens.">Influence of Specimen Geometry on the Estimation of the Planar Biaxial Mechanical Properties of Cruciform Specimens.</a> Exp. Mech. 54, 615-631 (2014).</li><li>Munarin, F., Kaiser, N. J., Kim, T. Y., Choi, B. R., Coulombe, K. L. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laser-Etched+Designs+for+Molding+Hydrogel-Based+Engineered+Tissues.">Laser-Etched Designs for Molding Hydrogel-Based Engineered Tissues.</a> Tissue Eng. Part C Methods. 23, 311-321 (2017).</li><li>Zhang, H., Chiao, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anti-fouling+Coatings+of+Poly(dimethylsiloxane)+Devices+for+Biological+and+Biomedical+Applications.">Anti-fouling Coatings of Poly(dimethylsiloxane) Devices for Biological and Biomedical Applications.</a> J. Med. Biol. Eng. 35, 143-155 (2015).</li></ol>

저자는 공개 없다.

조직 문화와 호환 되는 사용자 지정 된 PDMS 몰드 형상 조정 셀 정렬, 확산 속도, 효과적인 강성 같은 중요 한 설계 조직 속성에 훌륭한 유틸리티가 있다. 또한, 이러한 금형은 티슈를 준비 하 고 있는 기하학은 기계적 테스트16,17같은 중요 한 분석 응용 프로그램에 대 한 매우 유용 합니다. 전통적인 소와 관련 된 비용과 시간에 비해 할 때 특히 이러한 도?...

지난 2 년간 소프트 리소 그래피 사용 되었습니다 제조 기법으로 광범위 하 게 마이크로, 재료 연구, 및 조직 공학1,2의 분야에서 특히 과학 연구를 지원 하기 위해 3. 네거티브 마스터 몰드에서 만든은 원하는 모양으로 개체가 복제 성형 편리 하 고 저렴 한 비용의 방법 생산 긍정적인 PDMS는 복제 캐스팅 hydrogels 모양에 사용할 수 있습니다 제공 합니다. 그러나, 필요한 부정적인 마스터 금형은 일반적으로 비싼, 시간이 걸리는, 크기, 제한 된 제작 기술을 사용 하 여 생산과 클린 룸 공간 및 정교한 장비를 필요. 잠재적인 대안을 제공 하는 3D 인쇄, 자사의 유틸리티 저렴 프린터 및 일반적인 3D 프린터 고분자 및 치료 하는 것을 억제할 수 있는 PDMS 사이의 화학 상호 작용의 해상도 제한으로 인해 다소 제한 됩니다.
레이저 절단기 시스템 모두 절단 하 고 플라스틱, 나무, 유리, 그리고 금속 등 재료를 에칭 최근 획기적으로 저렴 하 고 따라서 더 연구 도구를 날조를 위한 접근 되고있다. 상업 학년 레이저 절단기 센티미터 규모 최소 기능 25 µ m 보다 작은 개체를 날조 할 수 있다 고 추가 필요 최소한의 교육, 전문 지식, 그리고 사용 하는 시간. PDMS의 레이저 절제 이전 마이크로 소자의 제조에 사용 되었습니다, 하는 동안 우리의 지식 없이 원고 프로세스는 밀리미터와 센티미터 규모 금형 레이저 부정적인 마스터 금형4 컷에서 날조 될 수 있다 설명 했다 .
영양 보급, 셀룰러 맞춤 및 기계적 성질5,,67을 개량 하기 위하여 설계 조직 형태를 조작 하는 주로이 방법을 사용 했습니다. 그러나,이 기술은 성형된 hydrogels 약물 전달 및 재료 과학 연구8등 관심 있는 모든 분야에서 활용에 대 한 수 있습니다. 레이저 커터에 대 한 액세스, PDMS 몰드 복제는 (그 것이이 논문의 범위를 벗어납니다 다중 파트 형 없이 제거 억제) 돌출부 없이 거의 모든 형상에 대 한 만들 수 있습니다 그리고 그 레이저 침대의 크기에 맞는.

<table><tbody><tr><td>&lt;strong&gt;Item&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Bovine fibrinogen</td><td>Sigma</td><td>F8630-5G</td><td>Constructs</td></tr><tr><td>Bovine thrombin</td><td>Sigma</td><td>T6634-250UN</td><td>Constructs</td></tr><tr><td>Bovine aprotinin</td><td>Sigma</td><td>10820-25MG</td><td>Constructs</td></tr><tr><td>Rat tail collagen I, 4 mg/mL</td><td>Advanced Biomatrix</td><td>5153-100MG</td><td>Constructs</td></tr><tr><td>Sodim chloride</td><td>Fisher</td><td>BP358-10</td><td>Constructs</td></tr><tr><td>PBS</td><td>Life Technologies</td><td>14190-250</td><td>Constructs</td></tr><tr><td>Fine forceps</td><td>Fine Science Tools</td><td>11252-20</td><td>Constructs</td></tr><tr><td>Sylgard 184 silicone elastomer</td><td>Corning</td><td>4019862</td><td>PDMS Molds</td></tr><tr><td>Lab tape</td><td>Fisher</td><td>15-901-5R</td><td>PDMS Molds</td></tr><tr><td>Acrylic, 1/4&quot; thick</td><td>McMaster-Carr</td><td>8560K356</td><td>PDMS Molds</td></tr><tr><td>HEPES Buffer, 1 M</td><td>Sigma</td><td>H3537-100ML</td><td>Constructs</td></tr><tr><td>RPMI 1640 medium, powder</td><td>Fisher</td><td>31800-089</td><td>Constructs</td></tr><tr><td>Calcium chloride dihydrate</td><td>Fisher</td><td>AC423520250</td><td>Constructs</td></tr><tr><td>Magnesium chloride hexahydrate</td><td>Fisher</td><td>M33 500</td><td>Constructs</td></tr><tr><td>Potassium chloride</td><td>Sigma</td><td>P9541-500G</td><td>Constructs</td></tr><tr><td>Sodium phosphate dibasic heptahydrate</td><td>Sigma</td><td>S9390-500G</td><td>Constructs</td></tr><tr><td>Glucose</td><td>Sigma</td><td>G5767-25G</td><td>Constructs</td></tr><tr><td>OCT</td><td>VWR</td><td>25608-930</td><td>Histology</td></tr><tr><td>Frozen block molds</td><td>VWR</td><td>25608-916</td><td>Histology</td></tr><tr><td>Hematoxylin</td><td>Fisher</td><td>3530 1</td><td>Histology</td></tr><tr><td>Eosin Y</td><td>Fisher</td><td>AC152880250</td><td>Histology</td></tr><tr><td>Fast green FCF</td><td>Fisher</td><td>AC410530250</td><td>Histology</td></tr><tr><td>&lt;strong&gt;Software&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Illustrator</td><td>Adobe Systems</td><td></td><td>Vector Graphics</td></tr><tr><td>Inkscape</td><td>(Open Source)</td><td></td><td>Vector Graphics</td></tr><tr><td>UCP (Universal Control Panel)</td><td>Universal Laser Systems</td><td></td><td>Laser Cutter Interface</td></tr><tr><td>&lt;strong&gt;Equipment&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>PLS6.75 Laser Cutter</td><td>Universal Laser Systems</td><td></td><td>Laser Cutter</td></tr><tr><td>Micromechanical Analyzer</td><td>Aurora Scientific</td><td>1530A with 5 mN load cell</td><td>Mechanical Analysis</td></tr></tbody></table>

custom engineered tissue culture molds from laser-etched masters

조직 공학의 분야는 계속 함 성숙에, 다양 한 조직 형태를 포함 한 조직 매개 변수에에서 대 한 관심 증가 되었습니다. 센티미터 규모 마이크로미터에 조직 형태를 조작 수 있습니다 직접 셀 정렬, 효과적인 기계적 특성을 변경 고 주소 영양 보급에 관련 된 제한 사항. 또한, 조직 준비는 선박 수 기계적 제약 조건을 셀 및 매트릭스 구조에 더 영향을 미칠 수 있는 스트레스 필드 결과로 조직에가 르 친다. 높은 재현성 크기와 모양의 조직 또한 생체 외에서 분석 실험을 샘플 크기는 전체 조직 기계적 분석 등 중요 한 유틸리티가 있다.
이 원고 레이저 에칭 아크릴에서 준비 하는 부정적인 마스터 금형을 이용 하는 다른 제조 방법에 설명 합니다: 이러한 금형입니다 (PDMS)와 함께 잘 수행, 센티미터 규모와 기능에 크기와 디자인을 허용 25 µ m 보다 작은 크기와 수 있습니다 빠르게 설계 및 조작 하는 최소한의 전문성과 저렴 한 비용. 최소한의 시간 및 비용 요구 사항을 빠르게 최적의 디자인을 결정 때까지 따라 반복 될을 넘어 조직 공학의 분야 들을 포함 한 관심의 모든 분석 결과 맞게 쉽게 적용할 레이저 에칭 금형에 대 한 허용.

레이저 절단기의 광학에 아주 약간 깊이 증가, 에칭으로 크기 감소 에칭된 영역 원인과 결과에 금형 레이저 광선의 가늘게 하 매우 미묘한 경사 벽으로 인해. 이 캐스트 PDMS 금형의 제거를 촉진 하는 데 도움이 됩니다 하지만 매우 깊이 부정적인 마스터 금형 에칭 경우 주의깊게 고려 되어야 한다 (> 6 m m)는 필수 (그림 1).
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Watch this Scientific Journal Video about Custom Engineered Tissue Culture Molds from Laser-etched Masters at JoVE.com

Custom Engineered Tissue Culture Molds from Laser-etched Masters

여기 우리는 복잡 한 형상의 설계 조직 히드로-기반 생산에 사용할 수 있는 사용자 지정입니다 금형 날조를 위한 신속 하 고, 손쉬운, 고 저가 방법 제시. 우리는 또한이 기술을 사용 하 여 생산 조작된 심장 조직에서 실시 하는 기계 및 조직학 평가 결과 설명 합니다.

사용자 지정 레이저 에칭 마스터스에서 조직 문화 금형 설계

As the field of tissue engineering has continued to mature, there has been increased interest in a wide range of tissue parameters, including tissue ...

custom-engineered-tissue-culture-molds-from-laser-etched-masters

Research

JoVE Journal

Bioengineering

6.4K 조회수.  Brown University. 여기 우리는 복잡 한 형상의 설계 조직 히드로-기반 생산에 사용할 수 있는 사용자 지정입니다 금형 날조를 위한 신속 하 고, 손쉬운, 고 저가 방법 제시. 우리는 또한이 기술을 사용 하 여 생산 조작된 심장 조직에서 실시 하는 기계 및 조직학 평가 결과 설명 합니다.

비디오: Custom Engineered Tissue Culture Molds from Laser-etched Masters

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.

기계 다이나믹 셀 문화 Microfabricated 플랫폼

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

연구

생체공학

Directed Cellular Self-Assembly to Fabricate Cell-Derived Tissue Rings for Biomechanical Analysis and Tissue Engineering

Each year, hundreds of thousands of patients undergo coronary artery bypass surgery in the United States.1 Approximately one third of these patients do not have suitable autologous donor vessels due to disease progression or previous harvest. The aim of vascular tissue engineering is to develop a suitable alternative source for these bypass grafts. In addition, engineered vascular tissue may prove valuable as living vascular models to study cardiovascular diseases. Several promising approaches to engineering blood vessels have been explored, with many recent studies focusing on development and analysis of cell-based methods.2-5 Herein, we present a method to rapidly self-assemble cells into 3D tissue rings that can be used in vitro to model vascular tissues.
To do this, suspensions of smooth muscle cells are seeded into round-bottomed annular agarose wells. The non-adhesive properties of the agarose allow the cells to settle, aggregate and contract around a post at the center of the well to form a cohesive tissue ring.6,7 These rings can be cultured for several days prior to harvesting for mechanical, physiological, biochemical, or histological analysis. We have shown that these cell-derived tissue rings yield at 100-500 kPa ultimate tensile strength8 which exceeds the value reported for other tissue engineered vascular constructs cultured for similar durations (&lt;30 kPa).9,10 Our results demonstrate that robust cell-derived vascular tissue ring generation can be achieved within a short time period, and offers the opportunity for direct and quantitative assessment of the contributions of cells and cell-derived matrix (CDM) to vascular tissue structure and function.

This article outlines a versatile method to create cell-derived tissue rings by cellular self-assembly. Smooth muscle cells seeded into ring-shaped agarose wells aggregate and contract to form robust three-dimensional (3D) tissues within 7 days. Millimeter-scale tissue rings are conducive to mechanical testing and serve as building blocks for tissue assembly.

자기 조립 셀룰러이 Biomechanical 분석 및 조직 공학을위한 셀 유래 조직 반지를 조작하는 감독

Each year, hundreds of thousands of patients undergo coronary artery bypass surgery in the United States.1 Approximately one third of these patients do ...

Evaluation of Cancer Stem Cell Migration Using Compartmentalizing Microfluidic Devices and Live Cell Imaging

In the last 40 years, the United States invested over 200 billion dollars on cancer research, resulting in only a 5% decrease in death rate. A major obstacle for improving patient outcomes is the poor understanding of mechanisms underlying cellular migration associated with aggressive cancer cell invasion, metastasis and therapeutic resistance1. Glioblastoma Multiforme (GBM), the most prevalent primary malignant adult brain tumor2, exemplifies this difficulty. Despite standard surgery, radiation and chemotherapies, patient median survival is only fifteen months, due to aggressive GBM infiltration into adjacent brain and rapid cancer recurrence2. The interactions of aberrant cell migratory mechanisms and the tumor microenvironment likely differentiate cancer from normal cells3. Therefore, improving therapeutic approaches for GBM require a better understanding of cancer cell migration mechanisms. Recent work suggests that a small subpopulation of cells within GBM, the brain tumor stem cell (BTSC), may be responsible for therapeutic resistance and recurrence. Mechanisms underlying BTSC migratory capacity are only starting to be characterized1,4.
Due to a limitation in visual inspection and geometrical manipulation, conventional migration assays5 are restricted to quantifying overall cell populations. In contrast, microfluidic devices permit single cell analysis because of compatibility with modern microscopy and control over micro-environment6-9.
We present a method for detailed characterization of BTSC migration using compartmentalizing microfluidic devices. These PDMS-made devices cast the tissue culture environment into three connected compartments: seeding chamber, receiving chamber and bridging microchannels. We tailored the device such that both chambers hold sufficient media to support viable BTSC for 4-5 days without media exchange. Highly mobile BTSCs initially introduced into the seeding chamber are isolated after migration though bridging microchannels to the parallel receiving chamber. This migration simulates cancer cellular spread through the interstitial spaces of the brain. The phase live images of cell morphology during migration are recorded over several days. Highly migratory BTSC can therefore be isolated, recultured, and analyzed further. 
Compartmentalizing microfluidics can be a versatile platform to study the migratory behavior of BTSCs and other cancer stem cells. By combining gradient generators, fluid handling, micro-electrodes and other microfluidic modules, these devices can also be used for drug screening and disease diagnosis6. Isolation of an aggressive subpopulation of migratory cells will enable studies of underlying molecular mechanisms.

A compartmentalizing microfluidic device for investigating cancer stem cell migration is described. This novel platform creates a viable cellular microenvironment and enables microscopic visualization of live cell locomotion. Highly motile cancer cells are isolated to study molecular mechanisms of aggressive infiltration, potentially leading to more effective future therapies.

Compartmentalizing 마이크로 유체 장치와 라이브 세포 이미징을 사용하여 암 줄기 세포 마이그레이션 평가

In the last 40 years, the United States invested over 200 billion dollars on cancer research, resulting in only a 5% decrease in death rate. A major ...

의학

Density Gradient Multilayered Polymerization (DGMP): A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Complex tissue culture matrices, in which types and concentrations of biological stimuli (e.g. growth factors, inhibitors, or small molecules) or matrix structure (e.g. composition, concentration, or stiffness of the matrix) vary over space, would enable a wide range of investigations concerning how these variables affect cell differentiation, migration, and other phenomena. The major challenge in creating layered matrices is maintaining the structural integrity of layer interfaces without diffusion of individual components from each layer1. Current methodologies to achieve this include photopatterning2-3, lithography4, sequential functionalization5, freeze drying6, microfluidics7, or centrifugation8, many of which require sophisticated instrumentation and technical skills. Others rely on sequential attachment of individual layers, which may lead to delamination of layers9. 
DGMP overcomes these issues by using an inert density modifier such as iodixanol to create layers of varying densities10. Since the density modifier can be mixed with any prepolymer or bioactive molecule, DGMP allows each scaffold layer to be customized. Simply varying the concentration of the density modifier prevents mixing of adjacent layers while they remain aqueous. Subsequent single step polymerization gives rise to a structurally continuous multilayered scaffold, in which each layer has distinct chemical and mechanical properties. The density modifier can be easily removed with sufficient rinsing without perturbation of the individual layers or their components. This technique is therefore well suited for creating hydrogels of various sizes, shapes, and materials.
A protocol for fabricating a 2D-polyethylene glycol (PEG) gel, in which alternating layers incorporate RGDS-350, is outlined below. We use PEG because it is biocompatible and inert. RGDS, a cell adhesion peptide11, is used to demonstrate spatial restriction of a biological cue, and the conjugation of a fluorophore (Alexa Fluor 350) enables us to visually distinguish various layers. This procedure can be adapted for other materials (e.g. collagen, hyaluronan, etc.) and can be extended to fabricate 3D gels with some modifications10.

Density Gradient Multilayered Polymerization DGMP: A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Here we describe a unique strategy for creating biocompatible, layered matrices with continuous interfaces between distinct layers for tissue engineering. Such a scaffold could provide an ideal customizable environment to modulate cell behavior by various biological, chemical or mechanical cues

밀도 그라데이션 Multilayered 중합 (DGMP) : 멀티 칸 만들기위한 소설 기법, 조직 공학을위한 사용자 정의 공사장 공중 발판

Complex tissue culture matrices, in which types and concentrations of biological stimuli (e.g. growth factors, inhibitors, or small molecules) or matrix ...

Microfabrication of Chip-sized Scaffolds for Three-dimensional Cell cultivation

Using microfabrication technologies is a prerequisite to create scaffolds of reproducible geometry and constant quality for three-dimensional cell cultivation. These technologies offer a wide spectrum of advantages not only for manufacturing but also for different applications. The size and shape of formed cell clusters can be influenced by the exact and reproducible architecture of the microfabricated scaffold and, therefore, the diffusion path length of nutrients and gases can be controlled.1 This is unquestionably a useful tool to prevent apoptosis and necrosis of cells due to an insufficient nutrient and gas supply or removal of cellular metabolites.
Our polymer chip, called CellChip, has the outer dimensions of 2 x 2 cm with a central microstructured area. This area is subdivided into an array of up to 1156 microcontainers with a typical dimension of 300 m edge length for the cubic design (cp- or cf-chip) or of 300 m diameter and depth for the round design (r-chip).2
So far, hot embossing or micro injection moulding (in combination with subsequent laborious machining of the parts) was used for the fabrication of the microstructured chips. Basically, micro injection moulding is one of the only polymer based replication techniques that, up to now, is capable for mass production of polymer microstructures.3 However, both techniques have certain unwanted limitations due to the processing of a viscous polymer melt with the generation of very thin walls or integrated through holes. In case of the CellChip, thin bottom layers are necessary to perforate the polymer and provide small pores of defined size to supply cells with culture medium e.g. by microfluidic perfusion of the containers.
In order to overcome these limitations and to reduce the manufacturing costs we have developed a new microtechnical approach on the basis of a down-scaled thermoforming process. For the manufacturing of highly porous and thin walled polymer chips, we use a combination of heavy ion irradiation, microthermoforming and track etching. In this so called "SMART" process (Substrate Modification And Replication by Thermoforming) thin polymer films are irradiated with energetic heavy projectiles of several hundred MeV introducing so-called "latent tracks" Subsequently, the film in a rubber elastic state is formed into three dimensional parts without modifying or annealing the tracks. After the forming process, selective chemical etching finally converts the tracks into cylindrical pores of adjustable diameter.

We present two processes for the microfabrication of porous polymer chips for three-dimensional cell cultivation. The first one is hot embossing combined with a solvent vapour welding process. The second one uses a recently developed microthermoforming process combined with ion track technology leading to a significant simplification of manufacture.

입체 세포 배양을위한 칩 크기의 공사장 공중 발판의 Microfabrication

Using microfabrication technologies is a prerequisite to create scaffolds of reproducible geometry and constant quality for three-dimensional cell ...

생물학

A Multi-compartment CNS Neuron-glia Co-culture Microfluidic Platform

We present a novel multi-compartment neuron co-culture microsystem platform for in vitro CNS axon-glia interaction research, capable of conducting up to six independent experiments in parallel for higher-throughput. We developed a new fabrication method to create microfluidic devices having both micro and macro scale structures within the same device through a single soft-lithography process, enabling mass fabrication with good repeatability.
The multi-compartment microfluidic co-culture platform is composed of one soma compartment for neurons and six axon/glia compartments for oligodendrocytes (OLs). The soma compartment and axon/glia compartments are connected by arrays of axon-guiding microchannels that function as physical barriers to confine neuronal soma in the soma compartment, while allowing axons to grow into axon/glia compartments. OLs loaded into axon/glia compartments can interact only with axons but not with neuronal soma or dendrites, enabling localized axon-glia interaction studies. The microchannels also enabled fluidic isolation between compartments, allowing six independent experiments to be conducted on a single device for higher throughput.
Soft-lithography using poly(dimethylsiloxane) (PDMS) is a commonly used technique in biomedical microdevices. Reservoirs on these devices are commonly defined by manual punching. Although simple, poor alignment and time consuming nature of the process makes this process not suitable when large numbers of reservoirs have to be repeatedly created. The newly developed method did not require manual punching of reservoirs, overcoming such limitations. First, seven reservoirs (depth: 3.5 mm) were made on a poly(methyl methacrylate) (PMMA) block using a micro-milling machine. Then, arrays of ridge microstructures, fabricated on a glass substrate, were hot-embossed against the PMMA block to define microchannels that connect the soma and axon/glia compartments. This process resulted in macro-scale reservoirs (3.5 mm) and micro-scale channels (2.5 &mu;m) to coincide within a single PMMA master. A PDMS replica that served as a mold master was obtained using soft-lithography and the final PDMS device was replicated from this master.
Primary neurons from E16-18 rats were loaded to the soma compartment and cultured for two weeks. After one week of cell culture, axons crossed microchannels and formed axonal only network layer inside axon/glia compartments. Axons grew uniformly throughout six axon/glia compartments and OLs from P1-2 rats were added to axon/glia compartments at 14 days in vitro for co-culture.

We developed a novel multi-compartment neuron co-culture microsystem platform for in vitro CNS axon-glia interaction research. The platform is capable of conducting up to six independent experiments in parallel and was fabricated using a newly developed macro/micro hybrid fabrication method.

다중 구획 CNS 뉴런 - glia 공동 문화 Microfluidic 플랫폼

We present a novel multi-compartment neuron co-culture microsystem platform for in vitro CNS axon-glia interaction research, capable of conducting up to ...

Fabrication of Custom Agarose Wells for Cell Seeding and Tissue Ring Self-assembly Using 3D-Printed Molds

Engineered tissues are being used clinically for tissue repair and replacement, and are being developed as tools for drug screening and human disease modeling. Self-assembled tissues offer advantages over scaffold-based tissue engineering, such as enhanced matrix deposition, strength, and function. However, there are few available methods for fabricating 3D tissues without seeding cells on or within a supporting scaffold. Previously, we developed a system for fabricating self-assembled tissue rings by seeding cells into non-adhesive agarose wells. A polydimethylsiloxane (PDMS) negative was first cast in a machined polycarbonate mold, and then agarose was gelled in the PDMS negative to create ring-shaped cell seeding wells. However, the versatility of this approach was limited by the resolution of the tools available for machining the polycarbonate mold. Here, we demonstrate that 3D-printed plastic can be used as an alternative to machined polycarbonate for fabricating PDMS negatives. The 3D-printed mold and revised mold design is simpler to use, inexpensive to produce, and requires significantly less agarose and PDMS per cell seeding well. We have demonstrated that the resulting agarose wells can be used to create self-assembled tissue rings with customized diameters from a variety of different cell types. Rings can then be used for mechanical, functional, and histological analysis, or for fabricating larger and more complex tubular tissues.

This protocol describes a platform for fabricating self-assembled tissue rings in variable sizes using a customized 3D-printed plastic mold. PDMS negatives are cured in the 3D-printed mold; then agarose is cast in the cured PDMS negatives. Cells are seeded into the resulting agarose wells where they aggregate into tissue rings.

셀 시드를 위한 사용자 지정 Agarose 우물 및 자기 조립 금형 3D 인쇄를 사용 하 여 조직 반지 제작

Engineered tissues are being used clinically for tissue repair and replacement, and are being developed as tools for drug screening and human disease ...

질병의 세포 상호 작용을 연구하기 위해 쥐 힘줄 집합체를 제작합니다.

Engineering Tendon Assembloids to Probe Cellular Crosstalk in Disease and Repair

Tendons enable locomotion by transferring muscle forces to bones. They rely on a tough tendon core comprising collagen fibers and stromal cell populations. This load-bearing core is encompassed, nourished, and repaired by a synovial-like tissue layer comprising the extrinsic tendon compartment. Despite this sophisticated design, tendon injuries are common, and clinical treatment still relies on physiotherapy and surgery. The limitations of available experimental model systems have slowed the development of novel disease-modifying treatments and relapse-preventing clinical regimes. 
In vivo human studies are limited to comparing healthy tendons to end-stage diseased or ruptured tissues sampled during repair surgery and do not allow the longitudinal study of the underlying tendon disease. In vivo animal models also present important limits regarding opaque physiological complexity, the ethical burden on the animals, and large economic costs associated with their use. Further, in vivo animal models are poorly suited to systematic probing of drugs and multicellular, multi-tissue interaction pathways. Simpler in vitro model systems have also fallen short. One major reason is a failure to adequately replicate the three-dimensional mechanical loading necessary to meaningfully study tendon cells and their function. 
The new 3D model system presented here alleviates some of these issues by exploiting murine tail tendon core explants. Importantly, these explants are easily accessible in large numbers from a single mouse, retain 3D in situ loading patterns at the cellular level, and feature an in vivo-like extracellular matrix. In this protocol, step-by-step instructions are given on how to augment tendon core explants with collagen hydrogels laden with muscle-derived endothelial cells, tendon-derived fibroblasts, and bone marrow-derived macrophages to substitute disease- and injury-activated cell populations within the extrinsic tendon compartment. It is demonstrated how the resulting tendon assembloids can be challenged mechanically or through defined microenvironmental stimuli to investigate emerging multicellular crosstalk during disease and injury.

저자 스포트라이트: 세포 메커니즘을 이해하기 위한 마우스 조립체를 개발하여 힘줄 연구 발전

Here, we present an assembloid model system to mimic tendon cellular crosstalk between the load-bearing tendon core tissue and an extrinsic compartment containing cell populations activated by disease and injury. As an important use case, we demonstrate how the system can be deployed to probe disease-relevant activation of extrinsic endothelial cells.

사용자 지정 레이저 에칭 마스터스에서 조직 문화 금형 설계

요약

더 많은 비디오 탐색

이 비디오의 챕터

기계 다이나믹 셀 문화 Microfabricated 플랫폼

자기 조립 셀룰러이 Biomechanical 분석 및 조직 공학을위한 셀 유래 조직 반지를 조작하는 감독

Compartmentalizing 마이크로 유체 장치와 라이브 세포 이미징을 사용하여 암 줄기 세포 마이그레이션 평가

밀도 그라데이션 Multilayered 중합 (DGMP) : 멀티 칸 만들기위한 소설 기법, 조직 공학을위한 사용자 정의 공사장 공중 발판

입체 세포 배양을위한 칩 크기의 공사장 공중 발판의 Microfabrication

다중 구획 CNS 뉴런 - glia 공동 문화 Microfluidic 플랫폼

셀 시드를 위한 사용자 지정 Agarose 우물 및 자기 조립 금형 3D 인쇄를 사용 하 여 조직 반지 제작

질병 및 복구에서 세포 누화를 조사하기 위한 힘줄 집합체 엔지니어링

질병 및 복구에서 세포 누화를 조사하기 위한 힘줄 집합체 엔지니어링

질병 및 복구에서 세포 누화를 조사하기 위한 힘줄 집합체 엔지니어링