Ortkrass Henning, Müller Marcel, Engdahl Anders Kokkvoll, Holst Gerhard, Huser Thomas
Biomolecular Photonics, Faculty of Physics, Bielefeld University, Bielefeld, Germany.
Excelitas PCO GmbH, Donaupark 11, Kelheim, Germany.
Nat Commun. 2024 Oct 15;15(1):8886. doi: 10.1038/s41467-024-53198-1.
High-resolution optical fluorescence microscopies and, in particular, super-resolution fluorescence microscopy, are rapidly adopting highly sensitive cameras as their preferred photodetectors. Camera-based parallel detection facilitates high-speed live cell imaging with the highest spatial resolution. Here, we show that the drive to use ever more sensitive, photon-counting image sensors in cameras can, however, have detrimental effects on the spatial resolution of the resulting images. This is particularly noticeable in applications that demand a high space-bandwidth product, where the image magnification is close to the Nyquist sampling limit of the sensor. Most scientists will often select image sensors based on parameters such as pixel size, quantum efficiency, signal-to-noise performance, dynamic range, and frame rate of the sensor. A parameter that is, however, typically overlooked is the sensor's modulation transfer function (MTF). We have determined the wavelength-specific MTF of front- and back-illuminated image sensors and evaluated how it affects the spatial resolution that can be achieved in high-resolution fluorescence microscopy modalities. We find significant differences in image sensor performance that cause the resulting spatial resolution to vary by up to 28%. This result shows that the choice of image sensor has a significant impact on the imaging performance of all camera-based optical microscopy modalities.
高分辨率光学荧光显微镜，尤其是超分辨率荧光显微镜，正迅速采用高灵敏度相机作为其首选的光电探测器。基于相机的并行检测有助于以最高空间分辨率进行高速活细胞成像。然而，我们发现，在相机中使用越来越灵敏的光子计数图像传感器的趋势，可能会对所得图像的空间分辨率产生不利影响。这在需要高空间带宽积的应用中尤为明显，即在图像放大倍数接近传感器的奈奎斯特采样极限的情况下。大多数科学家通常会根据像素大小、量子效率、信噪比性能、动态范围和传感器帧率等参数来选择图像传感器。然而，一个通常被忽视的参数是传感器的调制传递函数（MTF）。我们已经确定了前照式和背照式图像传感器的波长特异性MTF，并评估了它如何影响高分辨率荧光显微镜模式下可实现的空间分辨率。我们发现图像传感器性能存在显著差异，这导致最终的空间分辨率变化高达28%。这一结果表明，图像传感器的选择对所有基于相机的光学显微镜模式的成像性能都有重大影响。
