Schiffenbauer Yael S, Kalma Yael, Trubniykov Ella, Gal-Garber Orit, Weisz Lilach, Halamish Asaf, Sister Michael, Berke Gideon
Cell Kinetics, Lod, Israel.
Lab Chip. 2009 Oct 21;9(20):2965-72. doi: 10.1039/b904778f. Epub 2009 Jul 15.
Advances in molecular cell biology, medical research, and drug development are driving a growing need for technologies that enable imaging the dynamics of molecular and physiological processes simultaneously in numerous non-adherent living cells. Here we describe a platform technology and software--the CKChip system--that enables continuous, fluorescence-based imaging of thousands of individual living cells, each held at a given position ("address") on the chip. The system allows for sequential monitoring, manipulation and kinetic analyses of the effects of drugs, biological response modifiers and gene expression in both adherent and non-adherent cells held on the chip. Here we present four specific applications that demonstrate the utility of the system including monitoring kinetics of reactive oxygen species generation, assessing the intracellular enzymatic activity, measuring calcium flux and the dynamics of target cell killing induced by conjugated cytotoxic T-lymphocytes. We found large variations among individual cells in the overall amplitude of their response to stimuli, as well as in kinetic parameters such as time of onset, initial rate and decay of the response, and frequency and amplitude of oscillations. These variations probably reflect the heterogeneity of even cloned cell populations that would have gone undetected in bulk cell measurements. We demonstrate the utility of the system in providing kinetic parameters of complex cellular processes such as Ca++ influx, transients and oscillations in numerous individual cells. The CKChip opens up new opportunities in cell-based research, in particular for acquiring fluorescence-based, kinetic data from multiple, individual non-adherent cells.
分子细胞生物学、医学研究和药物开发的进展使得对相关技术的需求日益增长，这些技术能够在众多非贴壁活细胞中同时成像分子和生理过程的动态变化。在此，我们描述了一种平台技术和软件——CKChip系统，它能够对数千个单个活细胞进行基于荧光的连续成像，每个细胞都固定在芯片上的特定位置（“地址”）。该系统允许对芯片上贴壁和非贴壁细胞中药物、生物反应调节剂和基因表达的作用进行顺序监测、操作和动力学分析。在此，我们展示了四个具体应用，以证明该系统的实用性，包括监测活性氧生成的动力学、评估细胞内酶活性、测量钙通量以及偶联细胞毒性T淋巴细胞诱导的靶细胞杀伤动力学。我们发现，单个细胞对刺激的总体反应幅度以及诸如反应起始时间、初始速率和衰减、振荡频率和幅度等动力学参数存在很大差异。这些差异可能反映了即使是克隆细胞群体的异质性，而在大量细胞测量中这些差异可能未被发现。我们展示了该系统在提供复杂细胞过程（如众多单个细胞中的Ca++内流、瞬变和振荡）的动力学参数方面的实用性。CKChip为基于细胞的研究开辟了新机遇，特别是在从多个单个非贴壁细胞获取基于荧光的动力学数据方面。
