Sidelobe suppression in light-sheet fluorescence microscopy with Bessel beam plane illumination using subtractive imaging

The fluorescence from the out-of-focus region excited by the sidelobes of a Bessel beam is the major concern for light-sheet fluorescence microscopy (LSFM) with Bessel beam plane illumination. Here, we propose a method of applying the subtractive imaging to overcome the limitation of the conventional LSFM with Bessel beam plane illumination. In the proposed method, the sample is imaged twice by line scanning using the extended solid Bessel beam and the ring-like Bessel beam. By subtracting between the two images with similar out-of-focus blur, the improved image quality with the suppression of the Bessel beam sidelobes and enhanced sectioning ability with improved contrast are demonstrated.