Natan T. Shaked

Dual-interference-channel quantitative-phase microscopy of live cell dynamics

We introduce and experimentally demonstrate a fast and accurate method for quantitative imaging of the dynamics of live biological cells. Using a dual-channel interferometric setup, two phase-shifted interferograms of nearly transparent biological samples are acquired in a single digital camera exposure and digitally processed into the phase profile of the sample. Since two interferograms of the same sample are acquired simultaneously, most of the common phase noise is eliminated, enabling the visualization of millisecond-scale dynamic biological phenomena with subnanometer optical path length temporal stability.

Quantitative microscopy and nanoscopy of sickle red blood cells performed by wide field digital interferometry

We have applied wide-field digital interferometry (WFDI) to examine the morphology and dynamics of live red blood cells (RBCs) from individuals who suffer from sickle cell anemia (SCA), a genetic disorder that affects the structure and mechanical properties of RBCs. WFDI is a noncontact, label-free optical microscopy approach that can yield quantitative thickness profiles of RBCs and measurements of their membrane fluctuations at the nanometer scale reflecting their stiffness. We find that RBCs from individuals with SCA are significantly stiffer than those from a healthy control. Moreover, we show that the technique is sensitive enough to distinguish classes of RBCs in SCA, including sickle RBCs with apparently normal morphology, compared to the stiffer crescent-shaped sickle RBCs. We expect that this approach will be useful for diagnosis of SCA and for determining efficacy of therapeutic agents.

Generalized cell morphological parameters based on interferometric phase microscopy and their application to cell life cycle characterization

We present analysis tools which are formulated using wide-field interferometric phase microscopy measurements, and show their ability to uniquely quantify the life cycle of live cancer cells. These parameters are based directly on the optical path delay profile of the sample and do not necessitate decoupling the refractive index and the thickness in the cell interferometric phase profile, and thus can be calculated using a single-frame acquisition. To demonstrate the use of these parameters, we have constructed a wide-field interferometric phase microscopy setup and closely traced the full lifecycle of HeLa cancer cells. These initial results show the potential of the parameters to distinguish between the different phases of the cell lifecycle, as well others biological phenomena.

Two-step-only phase-shifting interferometry with optimized detector bandwidth for microscopy of live cells

We present a phase-shifting interferometric technique for imaging live biological cells in growth media, while optimizing spatial resolution and enabling potential real-time measurement capabilities. The technique uses slightly-off-axis interferometry which requires less detector bandwidth than traditional off-axis interferometry and fewer measurements than traditional on-axis interferometry. Experimental and theoretical comparisons between the proposed method and these traditional interferometric approaches are given. The method is experimentally demonstrated via phase microscopy of live human skin cancer cells.

Quantitative phase microscopy of biological samples using a portable interferometer

This Letter presents the τ interferometer, a portable and inexpensive device for obtaining spatial interferograms of microscopic biological samples without the strict stability and the highly coherent illumination that are usually required for interferometric microscopy setups. The device is built using off-the-shelf optical elements and can easily operate with low-coherence illumination, while being positioned in the output of a conventional inverted microscope. The interferograms are processed into the quantitative amplitude and phase profiles of the sample. Based on the phase profile, the optical-path-delay profile is obtained with temporal stability of 0.18 nm and spatial stability of 0.42 nm. Further experimental demonstration of using the τ interferometer for imaging the quantitative thickness profile of a live red blood cell is provided.

Compact and portable low-coherence interferometer with off-axis geometry for quantitative phase microscopy and nanoscopy

We present a simple-to-align, highly-portable interferometer, which is able to capture wide-field, off-axis interference patterns from transparent samples under low-coherence illumination. This small-dimensions and low-cost device can be connected to the output of a transmission microscope illuminated by a low-coherence source and measure sub-nanometric optical thickness changes in a label-free manner. In contrast to our previously published design, the τ interferometer, the new design is able to fully operate in an off-axis holographic geometry, where the interference fringes have high spatial frequency, and the interference area is limited only by the coherence length of the source, and thus it enables to easily obtain high-quality quantitative images of static and dynamic samples. We present several applications for the new design including nondestructive optical testing of transparent microscopic elements with nanometric thickness and live-cell imaging.

Review of three-dimensional holographic imaging by multiple-viewpoint-projection based methods

Methods of generating multiple viewpoint projection holograms of three-dimensional (3-D) realistic objects illuminated by incoherent white light are reviewed in this paper. Using these methods, it is possible to obtain holograms with a simple digital camera, operating in regular light conditions. Thus, most disadvantages characterizing conventional digital holography, namely the need for a powerful, highly coherent laser and extreme stability of the optical system, are avoided. The proposed holographic processes are composed of two stages. In the first stage, regular intensity-based images of the 3-D scene are captured from multiple points of view by a simple digital camera. In the second stage, the acquired projections are digitally processed to yield the complex digital hologram of the 3-D scene, where no interference is involved in the process. For highly reflecting 3-D objects, the resulting hologram is equivalent to an optical hologram of the objects recorded from the central point of view. We first review various methods to acquire the multiple viewpoint projections. These include the use of a microlens array and a macrolens array, as well as digitally generated projections that are not acquired optically. Next, we show how to digitally process the acquired projections to Fourier, Fresnel, and image holograms. Additionally, to obtain certain advantages over the known types of holograms, the proposed hybrid optical-digital process can yield novel types of holograms such as the modified Fresnel hologram and the protected correlation hologram. The prospective goal of these methods is to facilitate the design of a simple and portable digital holographic camera that can be useful for a variety of practical applications, including 3-D video acquisition and various types of biomedical imaging. We review several of these applications to signify the advantages of multiple viewpoint projection holography.

Whole-cell-analysis of live cardiomyocytes using wide-field interferometric phase microscopy.

We apply wide-field interferometric microscopy techniques to acquire quantitative phase profiles of ventricular cardiomyocytes in vitro during their rapid contraction with high temporal and spatial resolution. The whole-cell phase profiles are analyzed to yield valuable quantitative parameters characterizing the cell dynamics, without the need to decouple thickness from refractive index differences. Our experimental results verify that these new parameters can be used with wide field interferometric microscopy to discriminate the modulation of cardiomyocyte contraction dynamics due to temperature variation. To demonstrate the necessity of the proposed numerical analysis for cardiomyocytes, we present confocal dual-fluorescence-channel microscopy results which show that the rapid motion of the cell organelles during contraction preclude assuming a homogenous refractive index over the entire cell contents, or using multiple-exposure or scanning microscopy.

Tomographic phase microscopy with 180° rotation of live cells in suspension by holographic optical tweezers

We present a new tomographic phase microscopy (TPM) approach that allows capturing the three-dimensional refractive index structure of single cells in suspension without labeling, using 180° rotation of the cells. This is obtained by integrating an external off-axis interferometer for wide-field wave front acquisition with holographic optical tweezers (HOTs) for trapping and micro-rotation of the suspended cells. In contrast to existing TPM approaches for cell imaging, our approach does not require anchoring the sample to a rotating stage, nor is it limited in angular range as is the illumination rotation approach. Thus, it allows noninvasive TPM of suspended live cells in a wide angular range. The proposed technique is experimentally demonstrated by capturing the three-dimensional refractive index map of yeast cells, while collecting interferometric projections at an angular range of 180° with 5° steps. The interferometric projections are processed by both the filtered back-projection method and the diffraction theory method. The experimental system is integrated with a spinning disk confocal fluorescent microscope for validation of the label-free TPM results.

New procedures for minimizing the torque ripple in switched reluctance motors by optimizing the phase-current profile

This paper introduces five new optimization procedures for the minimization of the torque ripple in the switched reluctance motor (SRM). These new procedures are based on the optimization of the phase-current profile. Two optimization techniques, the simplex method and the genetic algorithm, are adapted to these optimization procedures. The paper compares an older optimization procedure, the optimum harmonic current injection procedure, and the new optimization procedure, and presents conclusions.