IDBR: CMOS cameras for high-frame-rate time-correlated single-photon counting

IDBR：用于高帧速率时间相关单光子计数的CMOS机生物成像技术的最新进展，特别是那些探索分子动力学的技术进步，正在超过技术创新。作为一种生物标志物，荧光寿命具有很大的潜力，可以揭示荧光团局部化学和物理环境的变化，以及通过激发态相互作用和Förster共振能量转移(FRET)来揭示单个蛋白质的结合动力学。许多最新的活性染料、分子探针甚至转基因标记策略都利用FRET来实时观察体外和体内的细胞过程。虽然FRET可以仅使用强度测量来检测，但定量可能会受到光漂白等实验因素的显著影响，而基于寿命的FRET测量明显更可靠。然而，荧光寿命成像显微镜(FLIM)在生物学研究中的采用和广泛使用受到两个主要因素的阻碍：获取FLIM图像的速度和FLIM所需的仪器的成本和复杂性。在这一多学科的方案中，将开发一种基于单光子雪崩二极管(SPAD)的新型二维高帧频互补金属氧化物半导体(CMOS)荧光寿命相机芯片。该芯片将应用于广域和基于激光扫描的显微技术，以实现电影成像方面的几个重要进展。在广域成像中，这将导致以高达1 kHz的入射光子限制帧速率采集图像。固态成像器主要基于两种技术，电荷耦合器件(CCD)和CMOS。这两种成像技术都是基于将光子转化为电子，并收集其中的许多电子来产生可测量的信号。这些成像器现在被用于各种类型的数码相机，从手机相机到用于生物成像的高端相机。由于光学技术在探测生物系统中如此普遍，相机代表着生物世界和固态世界之间的基本接口。在这项工作中，将设计一种全新的相机芯片，其基础是一种设备，它不是收集光子产生的电子，而是逐个对它们进行计数。这使得光子检测具有非常高的灵敏度。同时，它还可以分辨非常短(和很暗)的光学事件(约为10‘光秒的S)。这种能力将使新型生物成像应用成为可能。该项目支持对研究生和本科生进行多学科培训，并支持一项重要的K-12外联工作。

IDBR: CMOS cameras for high-frame-rate time-correlated single-photon countingRecent advances in biological imaging techniques, particularly those exploring molecular dynamics, are outpacing technological innovation. Fluorescence lifetime holds great potential as a biomarker that can reveal changes in a fluorophore's local chemical and physical environment, as well as the binding dynamics of single proteins through excited state interactions and Förster resonance energy transfer (FRET). Many of the latest active dyes, molecular probes and even transgenic labeling strategies exploit FRET to enable real-time observation of cellular processes both in-vitro and in-vivo. While FRET can be detected using intensity-only measurements, quantitation can be dramatically impaired by experimental factors such as photobleaching, whereas lifetime-based FRET measurements are significantly more robust. Nevertheless, adoption and widespread use of fluorescence lifetime imaging microscopy (FLIM) for biological research has been hindered by two major factors: the speed with which FLIM images can be acquired and the cost and complexity of the instrumentation required for FLIM. In this multidisciplinary proposal, a novel two-dimensional high-frame-rate complementary metal-oxide-semiconductor (CMOS) fluorescent lifetime camera chip based on single-photon avalanche diodes (SPADs) will be developed. This chip will be applied to both wide-field and laser-scanning-based microscopy techniques to enable several important advances in FLIM imaging. In widefield imaging, this will result in acquisition of images at a incident-photon-limited frame rate as high as 1 kHz.Solid-state imagers are based primarily on two technologies, charged-coupled device (CCD) and CMOS. Both of these imaging technologies are based on converting photons to electrons and collecting many of these electrons to produce a measurable signal. These imagers are now employed in digital cameras of every type, from cell phone cameras to the high-end cameras employed in biological imaging. Since optical techniques are so pervasive in probing biological systems, cameras represent the fundamental interface between the biological world and the solid-state world. In this effort, an entirely new camera chip will be designed based on a device that, instead of collecting electrons produced by photons, counts them, one-by-one. This enables very high sensitivity for photon detection. At the same time it allows resolution of very short (and dim) optical events (on the order of 10's of ps). Such capabilities will enable new types of biological imaging applications. This project supports the multidisciplinary training of graduate and undergraduate students and a significant K-12 outreach effort.