基于环形光瞳提高荧光辐射差分超分辨显微镜的成像分辨率

Improvement of Imaging Resolution of Fluorescence Emission Difference Super-Resolution Microscopy with Annular Pupil

荧光辐射差分(FED)显微术利用实心光斑扫描的一幅共聚焦图像减去空心光斑扫描的负共聚焦图像,实现了超分辨成像。使用简单的环形光瞳滤波器提高了无变形FED的成像分辨率。在空心焦斑光路上使用合适的环形光瞳滤波器,压缩空心焦斑的尺寸;同时,在实心焦斑的光路上允许使用更高数值孔径的孔径光阑,获得更小的、与空心焦斑相匹配的实心光斑分布:两方面作用下,提高了FED的空间分辨率。

Super-resolution imaging can be achieved by fluorescence emission difference(FED) microscopy through the subtraction of a negative confocal image scanned by a hollow focal spot from a confocal image scanned by a solid focal spot. This study proposes an easily accessible annular pupil filter to enhance the resolution of deformation-free FED imaging. The hollow spot size is reduced by an appropriate annular pupil filter in the light path of hollow spot. Meanwhile, a diaphragm with a high numerical aperture is permitted for the beam of a solid focal spot, and a small solid focal spot which is matching well with the hollow focal spot is achieved. The spatial resolution of FED microscopy is increased under the effects of these two aspects.