Fluorescence imaging of electrical activity in cardiac cells using an all-solid-state system
Fluorescence imaging of electrical activity in cardiac cells using an all-solid-state system
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DOI：
10.1109/tbme.2003.820376
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发表时间：
2004-02-01
影响因子：
4.6
通讯作者：
Tung, L
中科院分区：
文献类型：
--
作者：
Entcheva, E;Kostov, Y;Tung, L
在细胞水平上追踪心脏血管生成的空间和时间决定因素对所采用的光学标测技术提出了挑战。在本文中，我们描述了一个紧凑的系统，结合了两个非传统的低成本的解决方案，激发光源和发射滤波器的荧光测量跨膜电位，V-m，或细胞内钙，[Ca 2 +]（i）在心脏细胞网络。这是首次报道使用高功率蓝色和绿色发光二极管（LED）来激发用V-m-（di-8-ANEPPS）或[Ca 2 +]（i）-（Fluo-3）敏感染料染色的细胞单层。此外，我们使用简单的技术来制造合适的薄发射滤波器，其具有均匀的性质，没有自发荧光，高耐久性和良好的灵活性，用于成像V-m或[Ca 2 +]（i）。电池供电的LED和制造的发射滤波器，集成了光纤系统的接触荧光成像，被用作工具，表征在宏观尺度上的传导速度恢复。LED照明的多功能性通过以下方式得到进一步强调：1）在单细胞水平上展示其用于落射照明记录的用途，以及2）展示其独特的高频光调制能力。作为荧光测量的激发光源，LED显示出优异的稳定性;观察到可接受的信噪比和可忽略的细胞光损伤和指示剂染料光漂白。
Tracking spatial and temporal determinants of cardiac arrhythmogenesis at the cellular level presents challenges to the optical mapping techniques employed. In this paper, we describe a compact system combining two nontraditional low-cost solutions for excitation light sources and emission filters in fluorescence measurements of transmembrane potentials, V-m, or intracellular calcium, [Ca2+](i) in cardiac cell networks. This is the first reported use of high-power blue and green light emitting diodes (LEDs), to excite cell monolayers stained with V-m-(di-8-ANEPPS) or [Ca2+](i)-(Fluo-3) sensitive dyes. In addition, we use simple techniques for fabrication of suitable thin emission filters with uniform properties, no auto-fluorescence, high durability and good flexibility for imaging V-m or [Ca2+](i). The battery-operated LEDs and the fabricated emission filters, integrated with a fiber-optic system for contact fluorescence imaging, were used as tools to characterize conduction velocity restitution at the macro-scale. The versatility of the LEDs for illumination is further emphasized through 1) demonstration of their usage for epi-illumination recordings at the single-cell level, and 2) demonstration of their unique high-frequency light modulation ability. The LEDs showed excellent stability as excitation light sources for fluorescence measurements; acceptable signal-to-noise ratio and negligible cell photodamage and indicator dye photobleaching were observed.