We report on the generation of a hollow Bessel beam with a hole along the direction of propagation by using an easy-to-implement phase mask and investigate its effectiveness to reduce the out-of-focus background in light-sheet fluorescence microscopy (LSFM) with scanned Bessel beams by subtraction imaging. Overlaying ${\pi }$π-phase retardation between the two equal parts of the Bessel beam across the entrance pupil of the objective lens, a hollow Bessel beam with zero intensity at the focal plane can be achieved. By optimizing the numerical aperture of the annular mask applied in the hollow Bessel beam, matched distributions of the ring system between the hollow Bessel beam and the conventional Bessel beam are achieved. By subtraction between the two LSFM images, the out-of-focus blur caused by the ring system of the Bessel beam can be significantly reduced. Comparison with conventional Bessel LSFM images exhibits a better sectioning capability and higher contrast.

中文翻译：

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通过相位掩膜相减的方法可以增强扫描的贝塞尔光片显微镜的对比度。

我们通过使用易于实现的相位掩模报告了沿传播方向带有孔的空心贝塞尔光束的生成情况，并研究了其在光片荧光显微镜（LSFM）上减少散焦背景的有效性。通过减法成像扫描的贝塞尔光束。跨越物镜的入射光瞳的贝塞尔光束的两个相等部分之间的$ {\ pi} $π相位延迟重叠，可以实现在焦平面处具有零强度的空心贝塞尔光束。通过优化施加在空心贝塞尔光束中的环形掩模的数值孔径，可以实现环形系统在空心贝塞尔光束和常规贝塞尔光束之间的匹配分布。通过将两个LSFM图像相减，可以大大减少由贝塞尔光束的环形系统引起的离焦模糊。与传统的Bessel LSFM图像相比，具有更好的切片能力和更高的对比度。