Contact Fluorescence Imaging

描述（由申请人提供）：细胞结构和功能的光学荧光探针广泛用于生物学领域，如免疫学、组织化学、细胞凋亡、细胞粘附、趋化性、信号转导和电生理学。这些探针包括传统的基于化学的传感器和遗传编码的荧光蛋白生物传感器，所有这些都能够实时测量活细胞和组织中的生理信号。为此目的，冷却的电荷耦合器件（CCD）或光电二极管阵列通常与成像光学器件一起使用。这些商业系统具有有限的采样速度和成像区域尺寸的限制，并且通常体积大且成本高。一种新的方法，接触荧光成像（CFI），是一个简单的解决方案，在大面积的荧光信号的时空成像。这种方法的广泛应用面临的挑战是快速响应、具有高灵敏度的大幅面成像器的可用性。在探索R21阶段，我们将展示基于互补金属氧化物半导体（CMOS）技术的新型成像器的可行性。先进的模拟信号处理技术将使成像仪能够像素级基线减法，对比度模式操作，双波长成像，和比率成像，所有的能力，缺乏在商业上可用的成像阵列。在R33阶段，我们将扩大传感器阵列的尺寸，并生产一个可以从较小的模块化阵列平铺的大格式CMOS成像器。该项目的成果将是一个紧凑的成像系统，能够用荧光生物传感器监测静态和动态细胞事件。它将是微型的，便携式的，坚固的，可重复使用的，并适用于广泛的应用。我们将演示如何新的成像系统可以用来可视化的钙和电压波的时空动态在大尺寸的培养心肌细胞，以及其作为一个传统的荧光成像仪，可以连接到商业显微镜的相机端口的一般适用性。

DESCRIPTION (provided by applicant): Optical fluorescent probes of cellular structure and function are widely used in biology in fields as diverse as immunology, histochemistry, apoptosis, cell adhesion, chemotaxis, signal transduction, and electrophysiology. These probes include traditional chemically-based sensors and genetically-encoded fluorescent protein biosensors, all of which enable the measurement of physiological signals in real-time in living cells and tissue. For this purpose, cooled charge-coupled device (CCD) or photodiode arrays have typically been used with imaging optics. These commercial systems have limited sampling speed and limitations in the size of the imaging area, and are generally bulky and costly. A new method, Contact Fluorescence Imaging (CFI), is a simple solution for the spatiotemporal imaging of fluorescence signals over large areas. The challenge in terms of the widespread application of this method is the availability of a fast response, large format imager with high sensitivity. In the exploratory R21 phase, we will show feasibility of a new type of imager based on complementary metal oxide semiconductor (CMOS) technology. Advanced analog signal processing techniques will render the imager capable of pixel-level baseline subtraction, contrast mode operation, dual wavelength imaging, and ratiometric imaging, all capabilities that are lacking in commercially available imaging arrays. In the R33 phase, we will scale up the size of the sensor arrays, and produce a large format CMOS imager that can be tiled from smaller modular arrays. The outcome of this project will be a compact imaging system with the ability to monitor static and dynamic cellular events with fluorescent biosensors. It will be miniature, portable, rugged, reusable and adaptable to a wide range of applications. We will demonstrate how the new imaging system can be used to visualize the spatiotemporal dynamics of calcium and voltage waves in large-sized cultures of cardiac cells, as well as its general applicability as a conventional fluorescence imager that can be attached to the camera port of commercial microscopes.