Two-beam interference lattice lightsheet for structured illumination microscopy

Combining super-resolution structured illumination microscopy (SIM) and lattice lightsheet microscope (LLSM) has always been an idea approach for high spatiotemporal resolution over 3D application. We purpose a simpler method to perform 2D-SIM with 3 phases, which is 5/3 faster and less sensitive to the optical alignment compared to the 3D-SIM in LLSM. In this research, we modify the original square lattice lightsheet to become an ideal pattern for 2D-SIM by filtering the illumination pattern on the back pupil of the excitation objective. We show the generated lattice pattern is consistent in the experiment and the simulation. We achieved a spatial resolution of 184 ± 28 nm, 244 ± 48 nm, and 384 ± 20 nm in x, y, and z, respectively with 2D-SIM in LLSM, with exposure time 5 ms for each phase per plane. For biological applications, we perform 2D-SIM in LLSM by imaging the dynamics of actin and membrane ruffling in U2OS cell, with an exposure time of 20ms per phase with two colors recorded for 121 optical-sectioning planes per 3D stack.