Product Description
HyStem®-HP Hydrogel Kit - The growth factor delivery matrix
HyStem-HP hydrogel is fully chemically-defined and is ideal for cell applications whereby the slow, continuous release of growth factors is crucial to re-creating a desired microenvironment. The HyStem-HP Hydrogel Kit contains a combination of thiol-modified hyaluronan and a thiol-modified heparin (Heprasil®), thiol-modified denatured collagen (Gelin-S), and thiolreactive crosslinker, PEGDA (Extralink).The immobilized heparin in the HyStem-HP hydrogel mimics the heparin sulfate proteoglycans normally present in the extracellular matrix. Heparin forms an ionic bond with proteins and protects them from proteolysis and facilitates their slow release into the cell culture medium. This significantly reduces the amount of growth factor required to trigger cell growth or differentiation compared to when growth factors are added directly to the medium.Features
- Growth factors can be mixed into the hydrogels prior to gelation to provide a slow growth factor release depot.
- Hydrogels are suitable for animal implantation (such as angiogenesis applications and cell or drug delivery), culturing of primary cells, stem cells, and cell lines in the presence of growth factors.
- Cells can be encapsulated or grown on the hydrogel surface in any format, including culture flasks, 6- to 384-well plates or tissue culture inserts.
- Hydrogels can be easily customized by the user to possess the desired stiffness and gelation time by manipulating component concentration and mixing ratios.
GelationReconstituted HyStem-HP components remain liquid at 15 to 37°C. The hydrogel is formed when the crosslinking agent, Extralink®(PEGDA) is added to a mixture of Heprasil®(thiol-modified hyaluronan plus heparin) and Gelin-S®(thiol-modified gelatin). Gelation occurs in about twenty minutes after all three components are mixed. No steps depend on low temperatures or low pH. Diluting the components with phosphate-buffered saline (PBS) or cell-culture medium can increase the gelation time.3D Cell Recovery MatrixFor application where cell recovery is critical, the alternative crosslinker PEGSSDA is available for use with all HyStem, HyStem-C and HyStem-HP kits. This crosslinker provides the same advantages offered by Extralink with the additional benefit of containing easily reducible internal bonds. This allows for fast, easy recovery of single cells or clusters from the hydrogel for applications like RNA analysis or flow cytometry instead of slow enzymatic methods that can impact cell viability. Researchers are encouraged to contact us to determine the compatibility of particular cell types or culture systems with PEGSSDA.
Directions for Use
Download the HyStem®-Chydrogel kit instructions for:
Catalog #GS314 2.5 mL Trial Kit
Catalog #GS315 7.5 mL Kit
Catalog #GS1006 12.5 mL Kit
Product Q & A
Globular particles less than 75 kDa should be able to freely diffuse through a HyStem hydrogel.
When reconstituted using DG water, the pH of each HyStem component will be approximately 7.4-7.6.
One year from the date of receipt, if stored properly.
Any sterile, deionized, degassed water can be substituted for reconstitution. However, in order to ensure accurate and predictable dissolution and gelation times, our DG Water is highly recommended, as it is degassed, blanketed in argon, and has undergone validation testing with each HyStem component.
Gelin-S provides cellular attachment sites when incorporated in the hydrogel. Gelin-S is thiol-modified, denatured collagen I, derived from either bovine or porcine sources. Gelin-S is included in all HyStem-C and HyStem-HP kits.
Gelin-S has been thiol-modified in the same manner as the hyaluronan in Glycosil (or Heprasil), so that it covalently crosslinks with the Extralink in the HyStem hydrogels.
Yes. Peptides that contain a cysteine residue can be used. The cysteine residue must be present for the peptide to be covalently bonded to the hydrogel substrate.
Yes. ECM proteins, such as laminin, collagen, fibronectin, or vitronectin can be non-covalently incorporated into the hydrogel prior to crosslinking.
HyStem hydrogels and sponges differ in hydration and homogeneity. HyStem sponges are typically polymerized hydrogels that are subsequently freeze-dried. The resulting sponge is a fibrous, mesh network with pores and niches that enable cells to infiltrate and adhere. A true HyStem hydrogel is an encapsulating liquid that polymerizes around suspended cells in culture.
No. The compliance of the hydrogels is set by the amount of Extralink crosslinker added, the concentration of Glycosil (or Heprasil) and Gelin-S used, and the ratio of Glycosil (or Heprasil) to Gelin-S. Once this chemical structure of the hydrogel is fixed, it is not altered by prolonged exposure to cell culture medium.
HyStem sponges can be terminally sterilized by E-beam. HyStem hydrogels have not yet been validated for use with E-beam sterilization methods. HyStem hydrogels are not terminally sterilized by gamma irradiation.
Gelation time is affected by multiple aspects of the gel’s composition.One way to change the gelation time of a hydrogel is to vary the amount of crosslinker used. Gels with a lower amount of Extralink crosslinker will have a longer gelation time than those with a higher amount of crosslinker. Changing the amount of crosslinker will produce slight changes in gelation time.Gelation time can be dramatically changed by varying the Glycosil (or Heprasil) and Gelin-S concentrations. Concentrated solutions of Glycosil (or Heprasil) and Gelin-S will create a solution with a much shorter gelation time. This can easily be done by reconstituting the components in a smaller volume of DG Water. Alternatively, diluting these components in larger volumes of DG Water will dramatically increase the total time to form the hydrogel.
HyStem Hydrogels are virtually transparent and should not interfere with microscopy.
HyStem hydrogels may generate mild inflammation as part of the body’s natural healing process in response to injury. HyStem hydrogels do not trigger immune response when used in vivo. (These products are not for human use)
HyStem is degraded in vivo by matrix metalloproteinases (collagenases) and hyaluronidases.
Trypsin, Dipase, collagenase, and hyaluronidase have been used to help detach cells from the surface or from within HyStem hydrogels.
In general, the pore size for HyStem-C and HyStem-HP hydrogels is ~17 nm.
Product Applications
Click on the title of the desired protocol to learn more:
2D Cell Growth on HyStem Hydrogels
HyStem 3D Cell Encapsulation for Cell Delivery Applications Guide
HyStem 3D Cell Encapsulation in hydrogels using 96-well plates
HyStem 3D Cell Encapsulation in hydrogels using TC Inserts
Enzyme Digestion of HyStem Hydrogels for Recovery of Encapsulated Cells
Fluorescent Labeling of HyStem Hydrogels
Cell Recovery from Surface of HyStem Hydrogels
HyStem ECM Incorporation
HyStem Gelation Time Variation
HyStem Stiffness Variation Protocol for 7.5 mL kit
HyStem Stiffness Variation Protocol for 12.5 mL kit
Product References
References for HyStem®:
Gaetani, R., et al. (2015) Epicardial application of cardiac progenitor cells in a 3D-printed gelatin/hyaluronic acid patch preserves cardiac function after myocardial infarction. Biomaterials 61: 339-348.PMID: 17335875.Prestwich, G.D., et al. (2007) 3-D culture in synthetic extracellular matrices: new tissue models for drug toxicology and cancer drug discovery. Adv Enzyme Regul 47: 196-207.PMID: 17335875.Shu, X.Z., et al. (2006) Synthesis and evaluation of injectable, in situ crosslinkable synthetic extracellular matrices for tissue engineering. J Biomed Mater Res A 79: 901-912.PMID: 16941590.Shu, X.Z., et al. (2003) Disulfide-crosslinked hyaluronan-gelatin hydrogel films: a covalent mimic of the extracellular matrix for in vitro cell growth. Biomaterials 24: 3825-3834.PMID: 12818555.
S. Cai, et al. (2005)Injectable glycosaminoglycan hydrogels for controlled release of human basic fibroblast growth factor.Biomaterials, 26, 6054-6067.D. B. Pike, et al. (2006)Heparin-regulated release of growth factors in vitro and angiogenic response in vivo to implanted hyaluronan hydrogels containing VEGF and bFGF.Biomaterials, 27, 5242–5251.G. D. Prestwich, et al. (2007)3-D Culture in Synthetic Extracellular Matrices: New Tissue Models for Drug Toxicology and Cancer Drug Discovery.invited, Adv. Enz. Res., in press (2007).X. Z. Shu, et al, (2006)Synthesis and Evaluation of Injectable, In Situ Crosslinkable Synthetic Extracellular Matrices (sECMs) for Tissue Engineering.J. Biomed Mater. Res. A, 79A(4), 901-912.
Shu, X.Z., et al. (2004) In situ crosslinkable hyaluronan hydrogels for tissue engineering. Biomaterials 25: 1339-1348.PMID: 14643608.Mehra, T.D., et al. (2006) Molecular stenting with a crosslinked hyaluronan derivative inhibits collagen gel contraction. J Invest Dermatol 126: 2202-2209.PMID: 16741511.Shu, X.Z., et al. (2004) Attachment and spreading of fibroblasts on an RGD peptide-modified injectable hyaluronan hydrogel. J Biomed Mater Res A 68: 365-375.PMID: 14704979.Ghosh, K., et al. (2007) Cell adaptation to a physiologically relevant ECM mimic with different viscoelastic properties. Biomaterials 28: 671-679.PMID: 17049594.
Product Certificate of Analysis
Safety and Documentation
Certificate of Origin
Safety Data Sheet
Product Disclaimer
This product is for R&D use only and is not intended for human or other uses. Please consult the Material Safety Data Sheet for information regarding hazards and safe handling practices.
美国AdvancedBioMatrix（简称ABM） www.advancedbiomatrix.comAdvancedBioMatrix（简称ABM）是美国一家著名的生物公司，获得了AllerganInc的授权(Allergan用25年时间不断完善胶原蛋白相关的产品的生产工艺)，将Allergan的专业和技术用于蛋白生产与检测，致力于为组织工程、细胞分析及细胞增殖等研究领域提供优质稳定的产品。AdvancedBioMatrix不断丰富已有产品线，目前可为三维细胞培养提供各种胶原蛋白、纤连蛋白、玻连蛋白、水性凝胶、不同粘度与分子量的透明质酸以及低代成纤维细胞等。在美国全部产品授权Sigma销售。AdvancedBioMatrix是组织培养，细胞分析和细胞增殖三维（3D）应用的生命科学领域的领导者。我们的产品被公认为纯度，功能性和一致性的标准。我们在生产，分离，纯化，冷冻干燥，细胞培养和蛋白质测试，粘附肽，附着因子，底物刚性和其他3D矩阵产品方面拥有丰富的专业知识。我们的专业技术和知识正在被用来确保我们的产品质量最高，批次之间一致且易于为我们的研究客户使用。
美国AdvancedBioMatrix是3D组织培养、细胞检测和细胞增殖等领域实验解决方案的佼佼者。AdvancedBioMatrix在分离、纯化、冻干、细胞培养和蛋白检测、多肽粘附、附着因子、基质硬度和其他3Dmatrix 产品开发方面有着丰富的经验。AdvancedBioMatrix的研发经验和专业知识确保其产品可达到最佳质量，并保证产品之间一致性，方便研究客户使用。以下为AdvancedBioMatrix3DMatrices 产品竞争优势：1. 提供高纯度和成分确定的胞外基质；2. 超过1000余篇文献引用PureCol产品，品质非常均一；3. 在3D培养基领域可提供最全面的产品线；4. 唯一可提供特异性刚性有机硅基板的公司（CytoSoft）;5. 唯一可提供可溶性丝纤蛋白的供应商（可运用于多种3D培养）；6. 如果客户首次接触3D胶原凝胶，AdvancedBioMatrix还是唯一的预制胶原蛋白(PureColEZGel)供应商；
以下产品为AdvancedBioMatrix全球畅销品：1.PureCol 牛源I型胶原蛋白 3mg/ml#5005-100ML2.Nutragen牛源I型胶原蛋白 6mg/ml#5010-50ML3.FibriCol 牛源I型胶原蛋白 10mg/ml#5133-20ML4.VitroCol 人源I型胶原蛋白 #5007-20ML5. 弹性蛋白原 #5052-1MG6.ECMSelectArraykitUltra-36#5170-1EA7.CytoSoft（刚性可变的基底，AdvancedBioMatrix最新添加产品5190-7EA）8. 人III型胶原蛋白 #5021-10MG9. 人IV型胶原蛋白 #5022-5MG10.SilkFibroin溶液 #5154-20ML11.Fibronectin#5080-5MG12.Vitronectin#5051-0.1MG
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短截就是把枝条剪短，主要作用是促使其抽生新梢，增加分枝数目，以保证树势健状和正常结果。短截常用于骨干枝组修剪，结果枝组修剪，和树体局部更新复状。
短截按其长度可分为：
① 中短截：在一年生枝的中部短截，剪后萌发的顶端枝条，长势强，下部枝条长势弱。
② 重短截：剪去一年生枝的2/3。剪后萌发出的枝条较强状，一般用于主侧枝延长头修剪。
③ 重剪：剪去一年生枝的3/4－4/5，剪后萌发出的枝条长势强状，常用于发育枝作延长枝头和徒长果枝，中果枝的修剪。
④极重短截：剪去一年生枝的4/5以上，萌发后的枝条中庸偏状，常用于将发育枝和徒长枝培养结果枝组。
⑤留基部2芽剪：剪后萌发枝条较旺盛，常用于预备枝的修剪。对于幼龄树，树势较旺，以培养良好而牢固的树形结构，提早结果为主要目的，以轻短截，少疏间为主，从始果期到盛果期，主要使桃树多结果，并形成好的树形。
如果PCR产物不是很纯，或者PCR扩增条带比较小，PCR产物前面又有较多引物二聚体时，用胶回收，其余用PCR产物纯化试剂盒。
pcr纯化试剂盒和胶回收试剂盒的区别：
PCR纯化试剂盒：是直接水溶解的PAC产物就可以回收，回收效率高，但是只适合单一条带需要纯化测序的时候使用。
PCR凝胶试剂盒：是在PCR产物是混合物，有多条杂带的情况下，先跑胶将杂带分离，然后在将所要的条带位置的胶切下回收，后者的回收效率低，但是很纯净。
胶回收试剂盒操作步骤：
配制琼脂糖EB凝胶,电泳以分离DNA片段。任何类型或等级的琼脂糖都可以使用。
电泳足够时间后，在紫外灯下小心地把所需的DNA的片段切下来。并尽量去除多余的凝胶。
称取空离心管的重量，切下带目的片段的凝胶装在1.5ml离心管中并称其重量，求出凝胶块的重量，近似地确定其体积。一般情况下，凝胶的密度为1g/ml，于是凝胶的体积与重量的关系可按下面换算：凝胶薄片的重量为0.2g 则其体积为0.2ml；加入等倍凝胶体积的Binding Buffer，把混合物置于55℃~65℃水浴中温浴7min至凝胶完全融化，其间每隔2-3分钟混匀一次；
转移700μl的DNA-琼脂糖溶液到一个HiBindTM DNA柱子，并把柱子装在一个干净的2ml收集管内，室温下，10,000×g离心1min，弃去液体。
将柱子重新套回收集管中，加300μl Binding Buffer至HiBind DNA 柱子中；室温下，10,000×g离心 1分钟，去弃滤出液；这一步相当关键，不要忽略此步。
将柱子重新套回收集管中，加入700μl SPW Wash buffer至HiBind DNA柱子中，室温下，10,000×g离心1分钟，去弃滤出液；注：SPW Wash buffer在使用前必须按瓶子标鉴要求用无水乙醇进行稀释。
将柱子重新套回收集管中，重复加入700μl SPW Wash buffer至HiBind DNA柱子中，室温下，10,000×g离心1分钟，去弃滤出液；
弃去液体，将空柱子重新套回收集管中，10,000×g离心1min以甩干柱基质残余的液体。
这步可以去除柱子基质上残余的乙醇，不要省略此步―――对得到好的DNA产量是十分重要的。
把柱子装在一个干净的1.5ml离心管上，加入30~50μl洗脱液或灭菌水上柱子膜上，10,000×g离心1分钟，离心管中的溶液就是纯化的DNA产物，保存于-20度。
如果用组织DNA提取试剂盒，提出来的就是组织细胞的DNA了
过表达慢病毒
＞10^8 PFU/ml
干扰慢病毒
＞10^8 PFU/ml
