这项工作是从西弗吉尼亚大学玛丽巴布伦道夫癌症中心捐赠基金的支持。我们感谢“穆勒Susette（乔治城大学）和劳拉·凯利早期的意见和协助。西弗吉尼亚大学的显微成像设备的使用（支持玛丽巴布兰多夫癌症中心，国立卫生研究院资助P20 RR16440，P30 RR032138和P30 GM103488）表​​示感谢。

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没有利益冲突的声明。

辅助能力，可视化细胞降解细胞外基质中发现的早期步骤侵袭细胞的分子机制。陈文添在20世纪80年代初4,14,15首创，涂料荧光标记的细胞外蛋白质随后的微观分析的盖玻片上已经出现的主要技术评估侵袭伪足的功能范围广泛的细胞类型。规定的协议演示的基本的方法，用于制备明胶包被的盖玻片，形成一个小于2微米厚的胶原层适合于检测到的细胞外基质降解由细胞中最常规的荧光和激光?...

<table border="1"><tbody><tr><td> 试剂/仪器名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td>明胶</td><td>西格玛</td><td> G1890 </td><td>猪皮肤</td></tr><tr><td>蔗糖</td><td>费舍尔</td><td> BP220 </td><td></td></tr><tr><td> 12毫米盖玻片</td><td>费舍尔</td><td> 50-121-5159 </td><td></td></tr><tr><td> 24孔培养板</td><td>费舍尔</td><td> 08-772-1 </td><td> BD猎鹰</td></tr><tr><td>硝酸</td><td> Ricca </td><td> R5326000 </td><td> 20％的溶液</td></tr><tr><td>聚-L-赖氨酸</td><td>电子显微镜科学</td><td> 19320-B </td><td> 0.1％的溶液</td></tr><tr><td>戊二醛</td><td>西格玛</td><td> G7526 </td><td> 8％的溶液</td></tr><tr><td俄勒冈绿488-共轭明胶</td><td> Invitrogen公司</td><td> G-13186 </td><td></td></tr><tr><td>硼氢化钠</td><td>西格玛</td><td> 213462 </td><td></td></tr><tr><td>的Triton X-100的</td><td>费舍尔</td><td> BP151 </td><td></td></tr><tr><td>罗丹明 - 鬼笔环肽</td><td> Invitrogen公司</td><td> R415 </td><td></td></tr><tr><td>反皮层蛋白（克隆4F11） </td><td>密理博</td><td> 05-180 </td><td></td></tr><tr><td> ANTI-FLAG抗体</td><td>密理博</td><td> MAB3118 </td><td></td></tr><tr><td>的Alexa Fluor 647羊抗鼠IgG </td><td> Invitrogen公司</td><td> A21235 </td><td></td></tr><tr><td>延长Gold抗淬灭</td><td> Invitrogen公司</td><td> P36930 </td><td></td></tr><tr><td> LSM 510共聚焦显微镜</td><td>蔡司</td><td></td><td></td></tr><tr><td> ImageJ软件</td><td>公共领域</td><td></td><td> <a href="http://www.macbiophotonics.ca/imagej/" target="_blank">http://www.macbiophotonics.ca/imagej/</a> </td></tr><tr><td> AutoQuant X2.2软件</td><td>媒体控制论</td><td></td><td></td></tr><tr><td> NIS Elements软件</td><td>尼康</td><td></td><td></td></tr></tbody></table>

quantitative measurement of invadopodia-mediated extracellular matrix proteolysis in single and multicellular contexts

局部组织的细胞浸润到是一个重要的发展和平衡的过程。 malregulated入侵并随后细胞运动是多个病理过程，包括炎症，心血管疾病和肿瘤细胞转移1的特征。 focalized蛋白水解降解的胞外基质（ECM）的组件中的上皮细胞或内皮基底膜是启动细胞侵袭的一个关键步骤。在肿瘤细胞中，已确定， 在体外分析广泛ECM的降解是通过腹侧肌动蛋白富集膜凸结构称为侵袭伪足2,3。侵袭伪足形式在紧密合到的ECM，在那里它们通过的作用，基质金属蛋白酶（MMPs）中度ECM击穿。肿瘤细胞的能力，以形成侵袭伪足直接相关与标注侵入本地基质和相关的血管成分3。 _content“&gt;可视化的侵袭伪足介导的细胞外基质降解的细胞荧光显微镜用染料标记的基质蛋白涂在玻片上已经成为最流行 ​​的技术评估基质蛋白水解的程度和细胞侵袭能力4,5。在这里，我们描述一个版本产生荧光标记的玻片上，利用商业可用的俄勒冈绿-488明胶共轭的标准方法，这种方法很容易扩展到迅速产生大量的涂盖玻片，我们中经常遇到的一些常见的微观文物在此过程中，如何将这些可避免的。最后，我们使用现成的电脑软件，允许标记的明胶基质的降解介导的单个细胞和整个细胞群的量化描述的标准化方法。所描述的程序提供了准确和可重复的监测能力侵袭伪足活性，也可以作为一个平台，用于评估调制蛋白的表达或测试抗侵入性的胞外基质降解的化合物对单和多细胞设置的疗效。

Watch this Scientific Journal Video about Quantitative Measurement of Invadopodia-mediated Extracellular Matrix Proteolysis in Single and Multicellular Contexts at JoVE.com

Quantitative Measurement of Invadopodia-mediated Extracellular Matrix Proteolysis in Single and Multicellular Contexts

我们描述了典型的方法涂有荧光可视化侵袭伪足介导的基质降解明胶的生产显微镜的盖玻片。利用现有的软件的计算技术，提出了一个混合的人口和量化结果由单个细胞内的基质蛋白水解为多细胞群体，涵盖整个微观领域。

定量测量的侵袭伪足介导的细胞外基质蛋白水解单和多细胞的情况下

Cellular invasion into local tissues is a process important in development and homeostasis.  Malregulated invasion and subsequent cell movement is ...

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19.1K 次观看.  West Virginia University. 我们描述了典型的方法涂有荧光可视化侵袭伪足介导的基质降解明胶的生产显微镜的盖玻片。利用现有的软件的计算技术，提出了一个混合的人口和量化结果由单个细胞内的基质蛋白水解为多细胞群体，涵盖整个微观领域。