CIL:43415, Mus musculus, mammary adenocarcinoma. In Cell Image Library

DA3 cells expressing the fluorescent protein mCherry were grown to 90% confluence in wells of 2 cm diameter in DMEM supplemented with 10% heat-inactivated FCS (Gibco-BRL) and treated with or without the Met inhibitor PHA665752 (2.5 µM) for 2 hrs. A scratch was generated using a 200 µl tip, and the cells were incubated with or without 80 ng ml-1 Met-Hepatocyte Growth Factor/Scatter Factor (HGF/SF) and subjected to time lapse confocal laser scanning microscopy (CLSM-510, Zeiss, Germany) for approximately 26 hrs, with images taken at 14.5 min intervals. The position of each scratch was predefined, and a macro that repetitively positions the microscope on each point was executed. The acquired differential interference contrast (DIC) channel of the time-lapse sequence (shown here) was used for the multi-cellular analysis; the red fluorescence channel was exploited for single cell tracking.
See also: Zaritsky A, Natan S, Ben-Jacob E, Tsarfaty I (2012). Emergence of HGF/SF -induced Coordinated Cellular Motility. PLOS ONE 7(9): e44671

将表达荧光蛋白mCherry的DA3细胞接种于直径2 cm的培养孔中，在添加10%热灭活胎牛血清（FCS，Gibco-BRL）的杜氏改良伊格尔培养基（DMEM）内培养至细胞汇合度达90%。随后分别采用或不采用2.5 μM的Met抑制剂PHA665752处理细胞2小时。使用200 μL移液器枪头制造细胞划痕，将细胞分别置于添加或不添加80 ng·mL⁻¹ Met-肝细胞生长因子/分散因子（HGF/SF）的培养基中孵育，并使用德国蔡司公司生产的CLSM-510型延时共聚焦激光扫描显微镜（confocal laser scanning microscopy, CLSM）进行时长约26小时的成像采集，每14.5分钟获取一张图像。预先设定所有划痕的成像位点，并运行可循环将显微镜定位至各目标点位的宏程序。本延时序列的差分干涉对比度（differential interference contrast, DIC）通道图像被用于多细胞分析，红色荧光通道则用于单细胞追踪。

另见：Zaritsky A, Natan S, Ben-Jacob E, Tsarfaty I (2012). 《HGF/SF诱导的协调细胞运动的涌现》. PLOS ONE 7(9): e44671