A successful application of self-interference digital holographic microscopy in combination with a sample-rotation-based tomography module for three-dimensional (3-D) label-free quantitative live cell imaging with subcellular resolution is demonstrated. By means of implementation of a hollow optical fiber as the sample cuvette, the observation of living cells in different 3-D matrices is enabled. The fiber delivers a stable and accurate rotation of a cell or cell cluster, providing quantitative phase data for tomographic reconstruction of the 3-D refractive index distribution with an isotropic spatial resolution. We demonstrate that it is possible to clearly distinguish and quantitatively analyze several cells grouped in a "3-D cluster" as well as subcellular organelles like the nucleoli and local internal refractive index changes.

Holographic tomography (HT) is an advanced label-free optical microscopic imaging method used for biological studies. HT uses digital holographic microscopy to record the complex amplitudes of a biological sample as digital holograms and then numerically reconstruct the sample’s refractive index (RI) distribution in three dimensions. The RI values are a key parameter for label-free bio-examination, which correlate with metabolic activities and spatiotemporal distribution of biophysical parameters of cells and their internal organelles, tissues, and small-scale biological objects. This article provides insight on this rapidly growing HT field of research and its applications in biology. We present a review summary of the HT principle and highlight recent technical advancement in HT and its applications.

Off-axis holographic multiplexing involves capturing several complex wavefronts, each encoded into off-axis holograms with different interference fringe orientations, simultaneously, with a single camera acquisition. Thus, the multiplexed off-axis hologram can capture several wavefronts at once, where each one encodes different information from the sample, using the same number of pixels typically required for acquiring a single conventional off-axis hologram encoding only one sample wavefront. This gives rise to many possible applications, with focus on acquisition of dynamic samples, with hundreds of scientific papers already published in the last decade. These include field-of-view multiplexing, depth-of-field multiplexing, angular perspective multiplexing for tomographic phase microscopy for 3-D refractive index imaging, multiple wavelength multiplexing for multiwavelength phase unwrapping or for spectroscopy, performing super-resolution holographic imaging with synthetic aperture with simultaneous acquisition, holographic imaging of ultrafast events by encoding different temporal events into the parallel channels using laser pulses, measuring the Jones matrix and the birefringence of the sample from a single multiplexed hologram, and measuring several fluorescent microscopy channels and quantitative phase profiles together, among others. Each of the multiplexing techniques opens new perspectives for applying holography to efficiently measure challenging biological and metrological samples. Furthermore, even if the multiplexing is done digitally, off-axis holographic multiplexing is useful for rapid processing of the wavefront, for holographic compression, and for visualization purposes. Although each of these applications typically requires a different optical system or processing, they all share the same theoretical background. We therefore review the theory, various optical systems, applications, and perspectives of the field of off-axis holographic multiplexing, with the goal of stimulating its further development.

Due to incompleteness of input data inherent to Limited Angle Tomography (LAT), specific additional constraints are usually employed to suppress image artifacts. In this work we demonstrate a new two-stage regularization strategy, named Generalized Total Variation Iterative Constraint (GTVIC), dedicated to semi-piecewise-constant objects. It has been successfully applied as a supplementary module for two different reconstruction algorithms: an X-ray type solver and a diffraction-wise solver. Numerical tests performed on a detailed phantom of a biological cell under conical illumination pattern show significant reduction of axial blurring in the reconstructed refractive index distribution after GTVIC is added. Analogous results were obtained with experimental data.

As the 3D quantitative phase imaging (QPI) methods mature, their further development calls for reliable tools and methods to characterize and compare their metrological parameters. We use refractive index engineering during two-photon laser photolithography to fabricate a life-scale phantom of a biological cell with internal structures that mimic optical and structural properties of mammalian cells. After verification with a number of reference techniques, the phantom is used to characterize the performance of a limited-angle holographic tomography microscope.