Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, Maryland, USA.
Nat Protoc. 2010 Jul;5(7):1273-80. doi: 10.1038/nprot.2010.42. Epub 2010 Jun 17.
Programmed subcellular release is an in vitro technique for the quantitative study of cell detachment. The dynamics of cell contraction are measured by releasing cells from surfaces to which they are attached with spatial and temporal control. Release of subcellular regions of cells is achieved by plating cells on an electrode array created by standard microfabrication methods. The electrodes are then biochemically functionalized with an arginine-glycine-aspartic acid (RGD)-terminated thiol. Application of a voltage pulse results in electrochemical desorption of the RGD-terminated thiols, triggering cell detachment. This method allows for the study of the full cascade of events from detachment to subsequent subcellular reorganization. Fabrication of the electrode arrays may take 1-2 d. Preparation for experiments, including surface functionalization and cell plating, can be completed in 10 h. A series of cell release experiments on one device may last several hours.
可编程亚细胞释放是一种体外技术,用于定量研究细胞脱落。通过空间和时间控制将附着在表面上的细胞释放出来,可以测量细胞收缩的动态。通过使用标准微制造方法创建的电极阵列将细胞种植在电极上,可以实现细胞亚区室的释放。然后,通过将精氨酸-甘氨酸-天冬氨酸(RGD)末端硫醇进行生化功能化来修饰电极。施加电压脉冲会导致 RGD 末端硫醇电化学解吸,从而引发细胞脱落。该方法可用于研究从脱落到随后的亚细胞重排的整个级联事件。电极阵列的制造可能需要 1-2 天。实验准备工作,包括表面功能化和细胞种植,可在 10 小时内完成。在一个设备上进行一系列细胞释放实验可能需要数小时。