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基于宽场时间相关单光子计数的光片荧光寿命成像显微镜

Lightsheet fluorescence lifetime imaging microscopy with wide-field time-correlated single photon counting.

作者信息

Hirvonen Liisa M, Nedbal Jakub, Almutairi Norah, Phillips Thomas A, Becker Wolfgang, Conneely Thomas, Milnes James, Cox Susan, Stürzenbaum Stephen, Suhling Klaus

机构信息

Randall Centre for Cell and Molecular Biophysics, King's College London, London, UK.

Department of Physics, King's College London, London, UK.

出版信息

J Biophotonics. 2020 Feb;13(2):e201960099. doi: 10.1002/jbio.201960099. Epub 2019 Nov 25.

Abstract

We report on wide-field time-correlated single photon counting (TCSPC)-based fluorescence lifetime imaging microscopy (FLIM) with lightsheet illumination. A pulsed diode laser is used for excitation, and a crossed delay line anode image intensifier, effectively a single-photon sensitive camera, is used to record the position and arrival time of the photons with picosecond time resolution, combining low illumination intensity of microwatts with wide-field data collection. We pair this detector with the lightsheet illumination technique, and apply it to 3D FLIM imaging of dye gradients in human cancer cell spheroids, and C. elegans.

摘要

我们报道了基于宽场时间相关单光子计数(TCSPC)的光片照明荧光寿命成像显微镜(FLIM)。使用脉冲二极管激光器进行激发,并使用交叉延迟线阳极图像增强器(实际上是一个单光子敏感相机)以皮秒时间分辨率记录光子的位置和到达时间,将微瓦级的低照明强度与宽场数据采集相结合。我们将该探测器与光片照明技术配对,并将其应用于人类癌细胞球状体和秀丽隐杆线虫中染料梯度的三维FLIM成像。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1391/7065631/220d109141b0/JBIO-13-e201960099-g001.jpg

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