Sidelobe-free deterministic 3D nanoscopy with λ/33 axial resolution.

Deterministic three-dimensional (3D) super-resolution microscopy can achieve light-matter interaction in a small volume, but usually with the axial extension distinctly more elongated than the lateral one. The isoSTED method combining two opposing objectives and multiple laser beams can offer high axial extension at λ/12 level, but at the cost of optical system complexity and inherent sidelobes. The high-order nonlinear effect by multiphoton excitation would benefit to achieve a sub-diffraction resolution as well as to suppress the sidelobes. Herein, to achieve an easy-to-use, sidelobe-free deterministic 3D nanoscopy with high axial resolution, we developed a purely physical deterministic strategy (UNEx-4Pi) by fusion of ultrahighly nonlinear excitation (UNEx) of photon avalanching nanoparticles and mirror-based bifocal vector field modulation (4Pi). The theoretical studies of UNEx-4Pi concept showed that the main peak of fluorescence spot became sharper and its large sidelobe height was suppressed with the increasing optical nonlinearity. In addition, the simplicity and robustness of UNEx-4Pi system were demonstrated utilizing a mirror-assisted single-objective bifocal self-interference strategy. Experimentally, UNEx-4Pi realized an extremely constringent focal spot without sidelobes observed, achieving an axial resolution up to λ/33 (26 nm) using one low-power CW beam. We also demonstrated the super-resolution ability of the UNEx-4Pi scheme to bioimaging and nuclear envelope of BSC-1 cells were stained and imaged at an axial resolution of 32 nm. The proposed UNEx-4Pi method will pave the way for achieving light-matter interaction in a highly confined space, thereby advancing cutting-edge technologies like deterministic super-resolution sensing, imaging, lithography, and data storage.