Fig. 1.

The IHLLS system. a) Schematics of the IHLLS systems with b) one diffractive lens of focal length, f_SLM = 400 mm at the phase shift θ₁ = 0, and c) two diffractive lenses with focal lengths f_d1 = 220 mm and f_d2 = 2356 mm, at the phase shift θ₁ = 0, superposed with a slight defocus to bring the objects in focus in the middle of the camera FOV; The system consists of a water immersed microscope objective MO (Nikon 25X, NA 1.1, WD 2 mm), lenses L₁=L₄ with focal lengths 175 mm, L₂=L₃ with focal lengths 100 mm; mirrors M₁, M₂, M₃; polarizer P; 520 center wavelength, 40 nm band pass filter BPF; spatial light modulator SLM. The light propagates through either pathway 1 (blue line in a) for the original LLS or pathway 2 (red line in a)) for IHLLS, depending on the orientation of sliding mirror. A collimated 30 Bessel beam is focused by an excitation objective lens (d, f) which generates a lattice light sheet. The vectors represent the x, y, z and s planes of the Bessel. z and x are moved by the z and x galvos. It excites only fluorophores in the focal plane (e) and in/off the focal plane (g) of the detection objective lens. While the z-galvo and z-piezo are moved along the z axis to acquire stacks in LLS (d-e), in IHLLS only the z-galvo is moved at various z positions (f-g), (Visualization 1 ^{(5.8MB, avi)} ). For IHLLS, the size of the beam coming out the objective is diminished in half by the relay lens system, L₁ and L₂, to fit the size of the SLM. The SLM plane is optically conjugated with the objective back-focal-plane. The diffraction mask was positioned for all experiments on the anulus of 0.55 outer NA and 0.48 inner NA. The CMOS camera, tube lens, filter, and detection objective lens are used for fluorescence detection. The detection magnification $M_{T-LLS}$=62.5 and the illumination wavelength ${\lambda }_{illumination}$ = 488 nm. The width of the light sheet in the center of the FOV is about 400 nm. x-axis is the direction of the x-galvo mirror motion, z-axis is the direction of the z-piezo mirror motion, and s-axis is the direction of excitation light propagation.