Keefe T. Chan, Christa L. Cortesi, Anna Huttenlocher (2011) CIL:25706, Rattus, mammary adenocarcinoma. CIL. Dataset

MTLn3 cell cotransfected with control siRNA and GFP-cortactin to visualize invadipodia dynamics. The corresponding FAK knockdown experiments are CIL:25705 and CIL:25704. Fluorescence imaging was performed using a 60×/1.40 oil objective on an inverted microscope (1X-70;Olympus) in a 37°C closed system as previously described. Glass-bottomed dishes (35 mm) were coated with 10 µg/ml fibronectin for 1 h at 37°C. Cells were plated in Ham’s F12 containing 5% FBS and 20 mM Hepes, pH 7.2, and were allowed to adhere for 3 h. Fluorescent images were collected using a cooled CCD camera (CoolSNAP FX; Hamamatsu Photonics) and captured into MetaVue every 1 min for 1 h. Time-lapse sequences from live fluorescence imaging were first subjected to high-pass filtration based on the water algorithm (Zamir et al., 1999) to remove diffuse background fluorescence. Video corresponds to Fig. 1 D. and supplemental video 1 in J Cell Biol. 2009 Apr 20;185(2):357-70. Epub 2009 Apr 13. Bar, 10 µm.

MTLn3细胞共转染控制siRNA和GFP-肌动蛋白结合蛋白以可视化侵袭性伪足的动态变化。相应的FAK敲低实验编号分别为CIL:25705和CIL:25704。采用60×/1.40油镜在倒置显微镜（1X-70；Olympus）中，在37°C封闭系统中进行荧光成像，具体操作方法如前所述。底部为玻璃的培养皿（35mm）在37°C下用10 µg/ml纤连蛋白包被1小时。细胞在含有5%胎牛血清和20 mM Hepes，pH 7.2的Ham’s F12培养基中接种，并允许其粘附3小时。利用冷却型CCD相机（CoolSNAP FX；Hamamatsu Photonics）收集荧光图像，并每分钟捕捉一次，持续1小时，图像存储于MetaVue中。来自活细胞荧光成像的时间序列首先基于水算法（Zamir et al., 1999）进行高通滤波，以消除扩散背景荧光。视频对应于图1 D以及《细胞》杂志2009年4月20日第185卷第2期第357-370页的补充视频1。图例，10 µm。