近日，英国MRC分子生物学实验室Manton, James D.团队研究了具有不妥协时空分辨率的多光谱活细胞成像。相关论文于2025年9月8日发表在《自然—光子学》杂志上。

多光谱成像是一种成熟的方法，可以将荧光成像中可选颜色的数量扩展到典型的三种或四种以上。然而，标准方法不适合活细胞成像，因为需要将光分离到许多光谱通道中，并且解混算法难以处理低信噪比数据。

研究组介绍了一种在活细胞中进行多光谱成像的方法，该方法包括迭代光谱分解算法和基于八通道相机的图像采集硬件。这使得以视频速率捕获的低信噪比数据集能够准确解混，同时保持衍射有限的空间分辨率。该研究团队将这种方法应用于商用旋转盘共聚焦显微镜和自制斜平面光片显微镜上，同时对一到七种光谱上不同的荧光团进行成像，同时对荧光蛋白离子和小分子染料进行成像。研究组进一步开发了用有机荧光团标记的蛋白质结合蛋白（迷你粘合剂），并将其与多光谱成像方法相结合，以研究内源性水平上细胞表面受体的内体运输。

附：英文原文

Title: Multispectral live-cell imaging with uncompromised spatiotemporal resolution

Author: Kumar, Akaash, McNally, Kerrie E., Zhang, Yuexuan, Haslett-Saunders, Alex, Wang, Xinru, Guillem-Marti, Jordi, Lee, David, Huang, Buwei, Stallinga, Sjoerd, Kay, Robert R., Baker, David, Derivery, Emmanuel, Manton, James D.

Issue&Volume: 2025-09-08

Abstract: Multispectral imaging is an established method to extend the number of colours usable in fluorescence imaging beyond the typical limit of three or four. However, standard approaches are poorly suited to live-cell imaging owing to the need to separate light into many spectral channels, and unmixing algorithms struggle with low signal-to-noise ratio data. Here we introduce an approach for multispectral imaging in live cells that comprises an iterative spectral unmixing algorithm and eight-channel camera-based image-acquisition hardware. This enables the accurate unmixing of low signal-to-noise ratio datasets captured at video rates, while maintaining diffraction-limited spatial resolution. We use this approach on a commercial spinning-disk confocal microscope and a home-built oblique-plane light-sheet microscope to image one to seven spectrally distinct fluorophore species simultaneously, using both fluorescent protein fusions and small-molecule dyes. We further develop protein-binding proteins (minibinders), labelled with organic fluorophores, and use these in combination with our multispectral imaging approach to study the endosomal trafficking of cell-surface receptors at endogenous levels.

DOI: 10.1038/s41566-025-01745-7

Source: https://www.nature.com/articles/s41566-025-01745-7