Spectrally resolved fluorescence lifetime imaging^1,2,3 and spatial multiplexing^1,4,5 have offered information content and collection-efficiency boosts in microscopy, but efficient implementations for macroscopic applications are still lacking. An imaging platform based on time-resolved structured light and hyperspectral single-pixel detection has been developed to perform quantitative macroscopic fluorescence lifetime imaging (MFLI) over a large field of view (FOV) and multiple spectral bands simultaneously. The system makes use of three digital micromirror device (DMD)-based spatial light modulators (SLMs) to generate spatial optical bases and reconstruct N by N images over 16 spectral channels with a time-resolved capability (∼40 ps temporal resolution) using fewer than N² optical measurements. We demonstrate the potential of this new imaging platform by quantitatively imaging near-infrared (NIR) Förster resonance energy transfer (FRET) both in vitro and in vivo. The technique is well suited for quantitative hyperspectral lifetime imaging with a high sensitivity and paves the way for many important biomedical applications.

光谱解析的荧光寿命成像^1,2,3和空间多路复用^1,4,5在显微镜下提供了信息内容和收集效率的提高，但是仍然缺乏用于宏观应用的有效实现。已经开发了基于时间分辨结构光和高光谱单像素检测的成像平台，可以在大视野（FOV）和多个光谱带上同时执行定量宏观荧光寿命成像（MFLI）。该系统使用三种数字微镜装置（DMD）的基础的空间光调制器（SLM）来生成空间光碱和重建Ñ通过Ñ在与时间分辨能力16个光谱通道图像（〜使用比减少了40个皮秒时间分辨率） ñ ^2次光学测量。我们通过在体外和体内对近红外（NIR）福斯特共振能量转移（FRET）进行定量成像，证明了该新成像平台的潜力。该技术非常适合于具有高灵敏度的定量高光谱寿命成像，并为许多重要的生物医学应用铺平了道路。