Isotropic differential phase contrast microscopy for quantitative phase bio-imaging.

Quantitative phase imaging (QPI) has been investigated to retrieve optical phase information of an object and applied to biological microscopy and related medical studies. In recent examples, differential phase contrast (DPC) microscopy can recover phase image of thin sample under multi-axis intensity measurements in wide-field scheme. Unlike conventional DPC, based on theoretical approach under partially coherent condition, we propose a new method to achieve isotropic differential phase contrast (iDPC) with high accuracy and stability for phase recovery in simple and high-speed fashion. The iDPC is simply implemented with a partially coherent microscopy and a programmable thin-film transistor (TFT) shield to digitally modulate structured illumination patterns for QPI. In this article, simulation results show consistency of our theoretical approach for iDPC under partial coherence. In addition, we further demonstrate experiments of quantitative phase images of a standard micro-lens array, as well as label-free live human cell samples.

Talbot multi-focal holographic fluorescence endoscopy for optically sectioned imaging.

A wide-field multi-plane endoscopic system incorporating multiplexed volume holographic gratings and Talbot illumination to simultaneously acquire optically sectioned fluorescence images of tissue structures from different depths is presented. The proposed endoscopic system is configured such that multiple Talbot-illumination planes occur inside a volumetric sample and serve as the input focal planes for the subsequent multiplexed volume holographic imaging gratings. We describe the design, implementation, and experimental data demonstrating this endoscopic system's ability to obtain optically sectioned multi-plane fluorescent images of tissue samples in wide-field fashion without scanning in lateral and axial directions.

In-line digital holographic imaging in volume holographic microscopy.

A dual-plane in-line digital holographic imaging method incorporating volume holographic microscopy (VHM) is presented to reconstruct objects in a single shot while eliminating zero-order and twin-image diffracted waves. The proposed imaging method is configured such that information from different axial planes is acquired simultaneously using multiplexed volume holographic imaging gratings, as used in VHM, and recorded as in-line holograms where the corresponding reference beams are generated in the fashion of Gabor's in-line holography. Unlike conventional VHM, which can take axial intensity information only at focal depths, the proposed method digitally reconstructs objects at any axial position. Further, we demonstrate the proposed imaging technique's ability to effectively eliminate zero-order and twin images for single-shot three-dimensional object reconstruction.

Speckle-based volume holographic microscopy for optically sectioned multi-plane fluorescent imaging.

Structured illumination microscopy has been widely used to reconstruct optically sectioned fluorescence images in wide-field fashion; however, it still requires axial scanning to obtain multiple depth information of a volumetric sample. In this paper, a new imaging scheme, called speckle-based volume holographic microscopy system, is presented. The proposed system incorporates volumetric speckle illumination and multiplexed volume holographic gratings to acquire multi-plane images with optical sectioning capability, without any axial scanning. We present the design, implementation, and experimental image data demonstrating the proposed system's ability to simultaneously obtain wide-field, optically sectioned, and multi-depth resolved images of fluorescently labeled microspheres and tissue structures.

Wigner analysis of three dimensional pupil with finite lateral aperture.

A three dimensional (3D) pupil is an optical element, most commonly implemented on a volume hologram, that processes the incident optical field on a 3D fashion. Here we analyze the diffraction properties of a 3D pupil with finite lateral aperture in the 4-f imaging system configuration, using the Wigner Distribution Function (WDF) formulation. Since 3D imaging pupil is finite in both lateral and longitudinal directions, the WDF of the volume holographic 4-f imager theoretically predicts distinct Bragg diffraction patterns in phase space. These result in asymmetric profiles of diffracted coherent point spread function between degenerate diffraction and Bragg diffraction, elucidating the fundamental performance of volume holographic imaging. Experimental measurements are also presented, confirming the theoretical predictions.

Talbot holographic illumination nonscanning (THIN) fluorescence microscopy.

Optical sectioning techniques offer the ability to acquire three-dimensional information from various organ tissues by discriminating between the desired in-focus and out-of-focus (background) signals. Alternative techniques to confocal, such as active structured illumination, exist for fast optically sectioned images, but they require individual axial planes to be imaged consecutively. In this article, an imaging technique (THIN), by utilizing active Talbot illumination in 3D and multiplexed holographic Bragg filters for depth discrimination, is demonstrated for imaging in vivo 3D biopsy without mechanical or optical axial scanning.

Phase-coded volume holographic gratings for spatial-spectral imaging filters.

We present a design of phase-contrast filters embedded in a three-dimensional pupil to form phase-coded volume holographic gratings (VHGs) for spatial-spectral imaging. The phase-coded VHG improves image contrast and results in strong filtering properties to acquire weak phase structures of an object. In addition, incorporated with in-plane angle multiplexing, the multiplexed phase-coded VHGs enable obtaining weak phase information from multiple depths of an object. We experimentally demonstrate the multiplexed phase-coded VHGs for spatial-spectral imaging to enhance unstained features of spatial-spectral images of human breast cancer cells.

Enhancing the sensitivity to scattering coefficient of the epithelium in a two-layered tissue model by oblique optical fibers: Monte Carlo study.

Diffuse reflectance spectroscopy has been applied to detect tissue absorption and scattering properties associated with dysplasia, which is a potential precursor of epithelial cancers. The ability of DRS techniques to detect dysplasia could be improved by enhancing the detection of optical properties of the thin epithelial layer where dysplasia occurs. We propose a beveled fiber bundle probe consisting of a source fiber and multiple detection fibers parallel to each other and oriented obliquely to the tissue surface and investigate the sensitivity of reflectance measured with the probe to optical properties of a two-layered normal oral mucosa model. A scalable Monte Carlo method is employed to speed up analyzing spatially resolved reflectance spectra. Results reveal that the oblique probe is more sensitive to epithelial scattering and less sensitive to both stromal absorption and scattering than conventional perpendicular fiber configuration. The clinical relevance of the enhanced sensitivity to epithelial scattering by the proposed probe is demonstrated by quantifying optical properties of the two-layered tissue model from simulated data. The average error of extracted epithelial scattering coefficient is 1.5% and 32% using the oblique and perpendicular probe, respectively. The errors in other optical properties are all below 10% using the oblique probe.