Name: biophysical assays to probe the mechanical properties of the interphase cell nucleus: substrate strain application and microneedle manipulation
Uploaded: 2011-09-14T12:00:00
Description: Watch this Scientific Journal Video about Biophysical Assays to Probe the Mechanical Properties of the Interphase Cell Nucleus: Substrate Strain Application and Microneedle Manipulation at JoVE.com

这项工作是支持的美国国立卫生（R01 HL082792和R01 NS059348）和布里格姆与妇女医院心血管领导小组奖研究院。

1。基板应变的应用归核应变的测量包括应变菜的准备，透明，弹性硅膜细胞培养表面镀上的菜的细胞，获得细胞的图像（单轴或双轴）应变应用程序之前，期间和之后，。 制备硅膜的菜肴和细​​胞黏附<ol><li>每一株菜由一个特制的直径为3的无底的塑料盘“和一个塑料O型圈举行的硅膜，细胞培养基材，准备应变菜，钳4”X 4“一块硅膜之间的O形圈和菜。...

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	<li>Friedl, P., Wolf, K., Lammerding, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nuclear+mechanics+during+cell+migration.">Nuclear mechanics during cell migration.</a> Curr Opin Cell Biol. 23, 55-64 (2011).</li><li>Mejat, A., Misteli, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=LINC+complexes+in+health+and+disease.">LINC complexes in health and disease.</a> Nucleus. 1, 40-52 (2010).</li><li>Worman, H. J., Fong, L. G., Muchir, A., Young, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laminopathies+and+the+long+strange+trip+from+basic+cell+biology+to+therapy.">Laminopathies and the long strange trip from basic cell biology to therapy.</a> J Clin Invest. 119, 1825-1836 (2009).</li><li>Caille, N., Tardy, Y., Meister, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+strain+field+in+endothelial+cells+subjected+to+uniaxial+deformation+of+their+substrate.">Assessment of strain field in endothelial cells subjected to uniaxial deformation of their substrate.</a> Ann Biomed Eng. 26, 409-416 (1998).</li><li>Lammerding, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lamins+A+and+C+but+not+lamin+B1+regulate+nuclear+mechanics.">Lamins A and C but not lamin B1 regulate nuclear mechanics.</a> J Biol Chem. 281, 25768-25780 (2006).</li><li>Lammerding, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Abnormal+nuclear+shape+and+impaired+mechanotransduction+in+emerin-deficient+cells.">Abnormal nuclear shape and impaired mechanotransduction in emerin-deficient cells.</a> J Cell Biol. 170, 781-791 (2005).</li><li>Lammerding, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lamin+A/C+deficiency+causes+defective+nuclear+mechanics+and+mechanotransduction.">Lamin A/C deficiency causes defective nuclear mechanics and mechanotransduction.</a> J Clin Invest. 113, 370-378 (2004).</li><li>Verstraeten, V. L., Ji, J. Y., Cummings, K. S., Lee, R. T., Lammerding, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+mechanosensitivity+and+nuclear+stiffness+in+Hutchinson-Gilford+progeria+cells:+effects+of+farnesyltransferase+inhibitors.">Increased mechanosensitivity and nuclear stiffness in Hutchinson-Gilford progeria cells: effects of farnesyltransferase inhibitors.</a> Aging Cell. 7, 383-393 (2008).</li><li>Brooks, S. V., Zerba, E., Faulkner, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Injury+to+muscle+fibres+after+single+stretches+of+passive+and+maximally+stimulated+muscles+in+mice.">Injury to muscle fibres after single stretches of passive and maximally stimulated muscles in mice.</a> J Physiol. 488, 459-469 (1995).</li><li>Maniotis, A. J., Chen, C. S., Ingber, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Demonstration+of+mechanical+connections+between+integrins,+cytoskeletal+filaments,+and+nucleoplasm+that+stabilize+nuclear+structure.">Demonstration of mechanical connections between integrins, cytoskeletal filaments, and nucleoplasm that stabilize nuclear structure.</a> Proc Natl Acad Sci. 94, 849-854 (1997).</li></ol>

基板应变检测应变的应用我们和其他团体已经成功地用于研究诱导核细胞受机械应力的变形，并探讨具体的核衣壳蛋白的核刚度的贡献。4-8这种技术的优点是，它探针的机械性能生活在正常的细胞和细胞骨架的环境和核基板应变程序类似于生理负荷的应用程序，如发现在许多组织，如承包肌肉或血管壁。9此外，它使许多细胞的应变应用程序并行，增加的数量在?...

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td></tr><tr><td>纤维连接蛋白</td><td> Millipore公司</td><td> FC010 </td></tr><tr><td> MitoTracker红FM和绿色调频</td><td> Invitrogen公司</td><td> M22425和M - 7514 </td></tr><tr><td>赫希斯特33342 </td><td> Invitrogen公司</td><td> H3570 </td></tr><tr><td>汉克的缓冲盐生理盐水</td><td> Invitrogen公司</td><td> 14185 </td></tr><tr><td>苯酚自由，DMEM培养液</td><td> Invitrogen公司</td><td> 21063 </td></tr><tr><td>胎牛血清</td><td> Aleken生物</td><td> FBSS500 </td></tr><tr><td>青霉素/链霉素</td><td>西格玛</td><td> P0781 - 100ML </td></tr><tr><td>高硼硅玻璃与长丝</td><td>萨特仪器</td><td> BF100 - 78 - 10 </td></tr><tr><td>光泽度/光泽非增强硅薄膜，0.005“ </td><td>专业制造公司</td><td></td></tr><tr><td>贝科的磷酸盐缓冲液</td><td> Invitrogen公司</td><td> 14200 </td></tr><tr><td> 35毫米的玻璃底培养皿（FluoroDish） </td><td>世界精密仪器公司</td><td> FD35 - 100 </td></tr><tr><td> Braycote 804真空润滑脂</td><td>灵宝用品</td><td> 05133A - AB </td></tr></tbody></table>

biophysical assays to probe the mechanical properties of the interphase cell nucleus: substrate strain application and microneedle manipulation

在大多数真核细胞，细胞核是最大的细胞器，通常是2到10倍，比周围的细胞骨架的围追堵截，因此，原子核的物理性质，化学性质显着促进细胞生理和病理条件下的整体生物力学行为。例如，在中性粒细胞迁移和入侵的癌细胞，核刚度可以在外渗或通过狭窄的空间内组织的流逝带来的一大障碍。另一方面，机械活跃的组织，如肌肉细胞的细胞核，需要足够的结构支撑承受重复性的机械应力。重要的是，原子核是紧密集成到细胞结构，它是物理上连接到周围的骨架，这是关键的一个细胞核的细胞内运动和定位的要求，例如，在极化的细胞，在神经肌肉接头的突触核，或在细胞迁移。毫不奇怪，如lamins和nesprins，从而起到了关键作用，在确定核刚度和核苷酸骨架耦合的核衣壳蛋白的基因突变，已显示最近导致的人类疾病，包括金刚砂德赖富斯肌营养不良，肢带型肌营养不良症，和扩张型心肌病。探讨生物物理功能多样的核衣壳蛋白和特定的突变的影响，我们已开发的实验方法来研究原子核的物理性质，化学性质，在单，活细胞受到全球或局部的机械扰动。正是适用于基板应变的应用测量诱发核变形，细胞核变形产生重要的信息，并允许不同的突变或缺乏具体的核衣壳蛋白的细胞株之间的定量比较。用微针骨架应变本地化应用程序，是用来补充这个实验中，细胞内的细胞核和细胞骨架的之间的传力可以产生更多的信息。完整的活体细胞的核力学研究，保持正常的细胞内结构和避免潜在的孤立核工作时可能出现的文物。此外，基板应变的应用程序提供了一个很好的模式，在肌肉细胞或其他组织（例如，血管平滑肌细胞暴露血管应变）所经历的生理应激。最后，而这些工具已经被开发，主要研究核力学，他们也可以被应用于研究细胞骨架蛋白和mechanotransduction信号的功能。

Watch this Scientific Journal Video about Biophysical Assays to Probe the Mechanical Properties of the Interphase Cell Nucleus: Substrate Strain Application and Microneedle Manipulation at JoVE.com

Biophysical Assays to Probe the Mechanical Properties of the Interphase Cell Nucleus: Substrate Strain Application and Microneedle Manipulation

我们现在两个独立的显微镜为基础的工具来衡量，在应对全球性或局部应变的应用单一，生活贴壁细胞诱导核和细胞骨架变形。这些技术被用于确定核刚度（即变形）和探测细胞内的细胞核和细胞骨架的之间的传力。

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In most eukaryotic cells, the nucleus is the largest organelle and is typically 2 to 10 times stiffer than the surrounding cytoskeleton; consequently, the ...

Research

JoVE Journal

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Mechanical Stimulation of Stem Cells Using Cyclic Uniaxial Strain

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The role of mechanical forces in the development and maintenance of biological tissues is well documented, including several mechanically regulated ...

科研

生物学

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漂白实验（FRAP和FLIP）来衡量生活胚胎干细胞的染色质的蛋白质动力学

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Free-living planarian flatworms have a long history of experimental usage owing to their remarkable regenerative abilities1. Small fragments excised from ...

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5-methylcytosine (5-mC) constitutes ~2-8% of the total cytosines in human genomic DNA and impacts a broad range of biological functions, including gene ...

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超分辨率成像细菌科机械

Bacterial cell division requires the coordinated  assembly of more than ten essential proteins at midcell1,2. Central  to this process is the formation of ...

Application of an In vitro DNA Protection Assay to Visualize Stress Mediation Properties of the Dps Protein

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应用的<em在体外</em&gt;保护DNA检测，以可视化应力调解属性的DPS蛋白质

Oxidative stress is an unavoidable byproduct of  aerobic life.  Molecular oxygen is  essential for terrestrial metabolism, but it also takes part in many ...

The Cell-based L-Glutathione Protection Assays to Study Endocytosis and Recycling of Plasma Membrane Proteins

Membrane trafficking involves transport of proteins from the plasma membrane to the cell interior (i.e. endocytosis) followed by trafficking to lysosomes ...

Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs

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Probe-based quantitative PCR (qPCR) is a favoured method for measuring transcript abundance, since it is one of the most sensitive detection methods that ...

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The eye lens is a transparent organ that refracts and focuses light to form a clear image on the retina. In humans, ciliary muscles contract to deform the ...