Transport of intensity diffraction tomography with non-interferometric synthetic aperture for three-dimensional label-free microscopy.

We present a new label-free three-dimensional (3D) microscopy technique, termed transport of intensity diffraction tomography with non-interferometric synthetic aperture (TIDT-NSA). Without resorting to interferometric detection, TIDT-NSA retrieves the 3D refractive index (RI) distribution of biological specimens from 3D intensity-only measurements at various illumination angles, allowing incoherent-diffraction-limited quantitative 3D phase-contrast imaging. The unique combination of z-scanning the sample with illumination angle diversity in TIDT-NSA provides strong defocus phase contrast and better optical sectioning capabilities suitable for high-resolution tomography of thick biological samples. Based on an off-the-shelf bright-field microscope with a programmable light-emitting-diode (LED) illumination source, TIDT-NSA achieves an imaging resolution of 206 nm laterally and 520 nm axially with a high-NA oil immersion objective. We validate the 3D RI tomographic imaging performance on various unlabeled fixed and live samples, including human breast cancer cell lines MCF-7, human hepatocyte carcinoma cell lines HepG2, mouse macrophage cell lines RAW 264.7, Caenorhabditis elegans (C. elegans), and live Henrietta Lacks (HeLa) cells. These results establish TIDT-NSA as a new non-interferometric approach to optical diffraction tomography and 3D label-free microscopy, permitting quantitative characterization of cell morphology and time-dependent subcellular changes for widespread biological and medical applications.

Single-exposure 3D label-free microscopy based on color-multiplexed intensity diffraction tomography.

We present a 3D label-free refractive index (RI) imaging technique based on single-exposure intensity diffraction tomography (sIDT) using a color-multiplexed illumination scheme. In our method, the chromatic light-emitting diodes (LEDs) corresponding R/G/B channels in an annular programmable ring provide oblique illumination geometry that precisely matches the objective's numerical aperture. A color intensity image encoding the scattering field of the specimen from different directions is captured, and monochromatic intensity images concerning three color channels are separated and then used to recover the 3D RI distribution of the object following the process of IDT. In addition, the axial chromatic dispersion of focal lengths at different wavelengths introduced by the chromatic aberration of the objective lens and the spatial position misalignment of the ring LED source in the imaging system's transfer functions modeling are both corrected to significantly reduce the artifacts in the slice-based deconvolution procedure for the reconstruction of 3D RI distribution. Experimental results on MCF-7, Spirulina algae, and living Caenorhabditis elegans samples demonstrate the reliable performance of the sIDT method in label-free, high-throughput, and real-time (∼24 fps) 3D volumetric biological imaging applications.