Abstract
We demonstrate a multimodal superresolution microscopy technique based on a phase masked excitation beam in combination with spatially filtered detection. The theoretical foundation for calculating the focus from a non-paraxial beam with an arbitrary azimuthally symmetric phase mask is presented for linear and two-photon excitation processes as well as the theoretical resolution limitations. Experimentally this technique is demonstrated using two-photon luminescence from 80 nm gold particle as well as two-photon fluorescence lifetime imaging of fluorescent polystyrene beads. Finally to illustrate the versatility of this technique we acquire two-photon fluorescence lifetime, two-photon luminescence, and second harmonic images of a mixture of fluorescent molecules and 80 nm gold particles with > 120 nm resolution (λ/7). Since this approach exclusively relies on engineering the excitation and collection volumes, it is suitable for a wide range of scanning-based microscopies.
Highlights
IntroductionOver the past few decades, significant advancements have been made to overcome the spatial resolution limitations of conventional optical microscopy in order to probe nano-scale materials
Over the past few decades, significant advancements have been made to overcome the spatial resolution limitations of conventional optical microscopy in order to probe nano-scale materials.The earliest example is near field scanning optical microscopy [1], which helped shift the interest of the microscopy community towards superresolution techniques
The pattern on the spatial light modulator (SLM) is imaged onto the back aperture of a high numerical aperture (1.4 NA) microscope objective using relay lenses and focused onto the sample that is raster-scanned through the focus to create images
Summary
Over the past few decades, significant advancements have been made to overcome the spatial resolution limitations of conventional optical microscopy in order to probe nano-scale materials. We demonstrate that phase masked excitation in combination with selective detection can be utilized as a highly versatile superresolution technique that is suitable for a wide range of optical signals. In this case the side lobes are removed using spatially filtered detection, which results in artifact-free superresolution images. To demonstrate the versatility of this approach we acquired second harmonic generation (SHG), two-photon fluorescence (TPF), and TPL lifetime images of the same sample with
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