High-content biological microscopy targets high-resolution imaging across large fields-of-view (FOVs). Recent works have demonstrated that computational imaging can provide efficient solutions for high-content microscopy. Here, we use speckle structured illumination microscopy (SIM) as a robust and cost-effective solution for high-content fluorescence microscopy with simultaneous high-content quantitative phase (QP). This multi-modal compatibility is essential for studies requiring cross-correlative biological analysis. Our method uses laterally-translated Scotch tape to generate high-resolution speckle illumination patterns across a large FOV. Custom optimization algorithms then jointly reconstruct the samples super-resolution fluorescent (incoherent) and QP (coherent) distributions, while digitally correcting for system imperfections such as unknown speckle illumination patterns, system aberrations and pattern translations. Beyond previous linear SIM works, we achieve resolution gains of 4× the objectives diffraction-limited native resolution, resulting in 700 nm fluorescence and 1.2 μm QP resolution, across a FOV of 2 × 2.7 mm 2 , giving a space-bandwidth product (SBP) of 60 megapixels.

High-content biological microscopy targets high-resolution imaging across large fields-of-view, often achieved by computational imaging approaches. Previously, we demonstrated 2D multimodal high-content microscopy via structured illumination microscopy (SIM) with resolution > 2 × the diffraction limit, using speckle illumination from Scotch tape. In this work, we extend the method to 3D by leveraging the fact that the speckle illumination is in fact a 3D structured pattern. We use both a coherent and an incoherent imaging model to develop algorithms for joint retrieval of the 3D super-resolved fluorescent and complex-field distributions of the sample. Our reconstructed images resolve features beyond the physical diffraction-limit set by the systems objective and demonstrate 3D multimodal imaging with ∼ 0.6 × 0.6 × 6   μ m3 resolution over a volume of ∼ 314 × 500 × 24   μ m3.

Optical diffraction tomography (ODT) reconstructs a samples volumetric refractive index (RI) to create high-contrast, quantitative 3D visualizations of biological samples. However, standard implementations of ODT use interferometric systems, and so are sensitive to phase instabilities, complex mechanical design, and coherent noise. Furthermore, their reconstruction framework is typically limited to weakly scattering samples, and thus excludes a whole class of multiple-scattering samples. Here, we implement a new 3D RI microscopy technique that utilizes a computational multi-slice beam propagation method to invert the optical scattering process and reconstruct high-resolution (NA > 1.0) 3D RI distributions of multiple-scattering samples. The method acquires intensity-only measurements from different illumination angles and then solves a nonlinear optimization problem to recover the samples 3D RI distribution. We experimentally demonstrate the reconstruction of samples with varying amounts of multiple-scattering: a 3T3 fibroblast cell, a cluster of C. elegans embryos, and a whole C. elegans worm, with lateral and axial resolutions of ≤ 240 nm and ≤ 900 nm, respectively. The results of this work lays groundwork for future studies into using optical wavelengths to probe 3D RI distributions of highly scattering biological organisms.