Name: a quantitative evaluation of cell migration by the phagokinetic track motility assay
Uploaded: 2012-12-04T12:00:00
Description: Watch this Scientific Journal Video about A Quantitative Evaluation of Cell Migration by the Phagokinetic Track Motility Assay at JoVE.com

这项工作是由美国国家卫生研究院（AI050677，HD-051998，GM103433），马尔科姆·费斯特心血管研究奖学金，和美国心脏协会博士前的奖学金（10PRE4200007）的补助支持。

1。明胶涂层的盖玻片的制备<ol><li>放置酸洗玻璃盖玻片（15毫米直径），在无菌塑料100mm皿（晚餐）。将每道菜8-9盖玻片，并确保他们不接触对方或双方的菜.. </li></ol> 注：盖玻片，针和镊子必须是无菌的，以消除可能的污染的微生物，以及内毒素，这将影响细胞功能，包括蠕动。 <ol start="2"><li>称量明胶粉末，并重新悬浮在去离子水中的粉末，使明胶溶液的最终...

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Soc. 11, 55-75 (1951).</li><li>Frens, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Particle+size+and+sol+stability+in+mental+colloids.">Particle size and sol stability in mental colloids.</a> Colloid &#38; Polymer Science. 250, 736-741 (1972).</li><li>Frens, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+nucleation+for+the+regulation+of+the+particle+size+in+monodisperse+gold+suspensions.">Controlled nucleation for the regulation of the particle size in monodisperse gold suspensions.</a> Phys. Sci. 241, 20-22 (1973).</li><li>Zetter, B. 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轨道活力的phagokinetic在这篇文章中提出的分析是定量分析细胞迁移的一个简单而非常有效的方法。由于多种细胞类型可以分析9-17，这种方法有可能跨越多个学科的广泛使用。胶体金镀膜玻璃盖玻片的使用允许测量由移动的小区的轨道区清零。该法可衡量的效果，不同的刺激（ 如生长因子，纯化的ECM配体，病毒，细菌）对细胞活力。此外，缺乏监测细胞迁移的要求，进行细胞固定，...

<table border="1"><tbody><tr><td> 的试剂的名称 </td><td> 公司 </td><td> 目录号 </td><td> 评论 </td></tr><tr><td>盖玻片（15毫米） </td><td> Fisher Scientific则</td><td> 12-545-83 </td><td></td></tr><tr><td>明胶300布鲁姆</td><td> Sigma-Aldrich公司</td><td> G-1890 </td><td></td></tr><tr><td>四氯金酸三水合物</td><td>费舍尔化工</td><td> G54-1 </td><td> 14.5毫米（最终工作液） </td></tr><tr><td>柠檬酸钠</td><td> Fisher Scientific则</td><td> BP327-500 </td><td> （最后工作溶液）0.5％ </td></tr><tr><td>多聚甲醛</td><td> Fisher Scientific则</td><td> O4042 </td><td> 3％（最后工作溶液） </td></tr><tr><td> 100毫米组织培养皿</td><td>萨尔斯塔特</td><td> 83.1802 </td><td></td></tr><tr><td> 12孔板</td><td> Fisher Scientific则</td><td> 08-772-29 </td><td></td></tr><tr><td> 24孔板</td><td> Fisher Scientific则</td><td> 07-200-84 </td><td></td></tr><tr><td>泰科的烤箱Hybridiser HB-1D </td><td> LabPlanet </td><td> 2040500 </td><td>标准的实验室烤箱就足够了</td></tr><tr><td> 10毫升血清学移液管</td><td>萨尔斯塔特</td><td> 86.1254.001 </td><td></td></tr><tr><td>吸取援助灌装机/饮水机</td><td>德拉蒙德</td><td> 13-681-15 </td><td></td></tr><tr><td> P200单通道手动移液器</td><td> RAININ </td><td> PR-200 </td><td></td></tr><tr><td> 200毫升障碍提示</td><td>中电</td><td> BT200 </td><td></td></tr><tr><td> ImageJ软件</td><td> <a href="http://rsb.info.nih.gov/ij/" target="_blank">HTT</a>p :/ / rsb.info.nih.gov的/ IJ / </td><td></td><td>许可证：公共领域</td></tr><tr><td>尼康的Eclipse TE300一个配光CoolSNAPfx的单色12位的CCD相机</td><td>尼康</td><td></td><td>停产;最具可比性的规格有尼康的Eclipse钛，以及较低的尼康80I将是适当的。其他品牌也提供了类似的显微镜。 </td></tr><tr><td></td><td></td><td></td><td> 注意：下面列出的试剂和设备已动用我们在我们的各种研究。其他供应品，供应商，试剂和设备都可以使用，只要他们有类似的规范。 </td></tr></tbody></table>

a quantitative evaluation of cell migration by the phagokinetic track motility assay

单细胞和多细胞生物体细胞的活力是一个重要的生物过程。它的来源的营养物或远离不适合的条件，以及在多细胞生物组织的发展，免疫监视和伤口愈合，只是提了几个角色1,2,3的单细胞生物向运动是必不可少的。撤销这个过程可能会导致严重的神经系统，心血管疾病和免疫系统疾病，以及加剧了肿瘤的形成和传播4,5。分子，肌动蛋白聚合和受体回收已被证明发挥了重要作用，在细胞的扩展名（片状伪足），即推动向前运动的细胞6,7,8。然而，许多生物细胞迁移的问题仍然没有答案。 在人类健康和疾病的细胞运动中的核心作用突出的重要性，了解具体的MEChanisms参与了这一过程，并作出准确的方法来评估细胞的运动，特别是重要。显微镜通常是用于可视化细胞的运动。然而，细胞移动相当缓慢，耗费资源的过程，需要昂贵的相机和软件创建时间的推移定量电影运动细胞的细胞迁移的定量测量。因此，进行定量测量细胞迁移，是符合成本效益的，不费力，而且采用了常见的实验室设备的能力是一个伟大的许多研究人员的需要。 phagokinetic轨道蠕动测定法利用能力的移动单元以清除金颗粒，从它的路径建立一个可测量的磁道上的胶体金涂覆的玻璃盖玻片9,10。使用的自由软件，可以评估多个音轨，每次治疗完成统计的要求。该试验可以用来评估许多类型的细胞，如癌细胞11,12，成纤维细胞9，嗜中性粒细胞13，骨骼肌细胞14，角质形成细胞15，的滋养层16，血管内皮细胞17，和单核细胞10,18-22能动性。该协议涉及创建由柠檬酸钠，氯金酸（互惠3 +）的减少所生成的金纳米粒子（互惠°）涂覆的幻灯片。这种方法的开发由Turkevich 等人于1951年23，然后在20世纪70年代的改善Frens 等。24,25。作为一个结果，该化学还原步骤中，金颗粒（10-20 nm直径）从反应混合物中沉淀，并可以应用到玻璃盖玻片，然后准备用于在细胞迁移分析9,26,27。 在一般情况下，phagokinetic轨道蠕动法是一种快速，定量和容易的细胞能动性的措施。此外，它可以用来作为一个简单的高通量的检测，使用的细胞类型，不适合时间推移的成像，以及其他用途，取决于需要的研究员。一起，定量地测量多种细胞类型的细胞运动，而不需要昂贵的显微镜和软件的能力，以及常见的实验室设备和化学品的使用，作出的phagokinetic的轨道活力测定了坚实的选择，有兴趣的科学家了解细胞蠕动。

所示出的是示出了轨道区清零由一个单一的细胞（单核细胞从我们的实验是在图2中所示）在光学显微镜下拍摄的图像的一个例子。非运动细胞创建特征的小，椭圆形或圆形大片自己周围指示低基础水平的运动为这些未刺激的细胞（ 图2A和2B）。高度运动细胞[在我们的系统中，人巨细胞病毒（HCMV）感染的细胞]与此相反，其特征在于由作为细长的轨道区?...

Watch this Scientific Journal Video about A Quantitative Evaluation of Cell Migration by the Phagokinetic Track Motility Assay at JoVE.com

A Quantitative Evaluation of Cell Migration by the Phagokinetic Track Motility Assay

phagokinetic蠕动轨道法是一种方法，用于评估细胞的运动。具体而言，测定测量化学增活现象（随机细胞运动性），以定量的方式随着时间的推移。该试验利用细胞的能力，以建立一个可测量其运动轨迹胶体金涂覆盖玻片。

细胞迁移的的Phagokinetic轨道运动MLCK的定量评价

Cellular motility is an important biological process for both unicellular and multicellular organisms. It is essential for movement of unicellular ...

Research

JoVE Journal

Immunology and Infection

Quantitative Assessment of Immune Cells in the Injured Spinal Cord Tissue by Flow Cytometry:  a Novel Use for a Cell Purification Method

免疫细胞在受伤的脊髓组织流式细胞仪定量评估：一个细胞的分离纯化方法的新颖使用

Detection of immune cells in the injured central nervous system (CNS) using morphological or histological techniques has not always provided true ...

科研

免疫与感染

Quantitative Analyses of all Influenza Type A Viral Hemagglutinins and Neuraminidases using Universal Antibodies in Simple Slot Blot Assays

所有流感的定量分析类型使用通用抗体的一种病毒Hemagglutinins和Neuraminidases，在简单的狭缝杂交分析

Hemagglutinin (HA) and neuraminidase (NA) are two surface proteins of influenza viruses which are known to play important roles in the viral life cycle ...

Detection of Infectious Virus from Field-collected Mosquitoes by Vero Cell Culture Assay

从现场收集到的蚊子传染病毒Vero细胞培养法检测

Mosquitoes transmit a number of distinct viruses including important human pathogens such as West Nile virus, dengue virus, and chickungunya virus. Many ...

Quantitative High-throughput Single-cell Cytotoxicity Assay For T Cells

定量高通量单细胞的细胞毒性T细胞的测定

Cancer immunotherapy can harness the specificity of  immune response to target and eliminate tumors. Adoptive cell therapy (ACT)  based on the adoptive ...

Monitoring Dendritic Cell Migration using 19F / 1H Magnetic Resonance Imaging

Monitoring Dendritic Cell Migration using 19F / 1H Magnetic Resonance Imaging

监测树突状细胞迁移<sup&gt; 19</sup&gt; F /<sup&gt; 1</sup&gt; H磁共振成像

Continuous  advancements in noninvasive imaging modalities such as magnetic resonance  imaging (MRI) have greatly improved our ability to study ...

In vitro Coculture Assay to Assess Pathogen Induced Neutrophil Trans-epithelial Migration

In vitro Coculture Assay to Assess Pathogen Induced Neutrophil Trans-epithelial Migration

体外 共生分析评估病原体诱导中性粒细胞跨上皮迁移

Mucosal surfaces serve as protective barriers against pathogenic organisms.&#160;Innate immune responses are activated upon sensing pathogen leading to ...

Quantitative Assessment of Human Neutrophil Migration Across a Cultured Bladder Epithelium

人中性粒细胞迁移的跨越培养膀胱上皮定量评价

The recruitment of immune cells from the periphery to the site of inflammation is an essential step in the innate immune response at any mucosal surface. ...

Conformational Evaluation of HIV-1 Trimeric Envelope Glycoproteins Using a Cell-based ELISA Assay

HIV-1的三聚体包膜糖蛋白的构象评估使用基于细胞的ELISA检测

HIV-1 envelope glycoproteins (Env) mediate viral entry into target cells and are essential to the infectious cycle. Understanding how those glycoproteins ...

High-throughput Quantitative Real-time RT-PCR Assay for Determining Expression Profiles of Types I and III Interferon Subtypes

高通量实时定量RT-PCR检测方法的确定I型和III型干扰素亚型的表达谱

Described in this report is a qRT-PCR assay for the analysis of seventeen human IFN subtypes in a 384-well plate format that incorporates highly specific ...

Qualitative and Quantitative Assays for Detection and Characterization of Protein Antimicrobials

定性和定量测定为检测和蛋白质抗菌药物的表征

Initial evaluations of large microbial libraries for potential producers of novel antimicrobial proteins require both qualitative and quantitative methods ...