Study on the simplified phase-shifting digital holographic microscopy

In this paper, a new simplified technique for effectively eliminating the zero order and the conjugate virtual image in digital holographic microcopy, which makes use of two-step phase-shifting method of just recording two holograms and an intensity image of object wave, is proposed. Meanwhile, combined with the principle of making full use of spatial bandwidth of the CCD sensor by in-line lens-less Fourier holographic recording geometry, the theory and experimental methods to increase the resolution of the reconstructed image in digital holography by using phase-shifting technique are detailedly analyzed. At end, the validity and availability of this technique has been demonstrated through the off-axis and in-line Fourier transform recording geometry. The study provides some theoretical and experimental guidance for the design and operation of a digital holographic microscopy system.     

Phase reconstruction in lensless digital in-line holographic microscopy

The phase reconstruction in a digital in-line holographic microscopy is compared using two numerical reconstruction methods. The first method uses one Fourier transform and second one uses three Fourier transforms. It is shown that the latter method gives improved object phase reconstruction as compared to the former.     

Polarization phase shifting in digital holographic microscopy

In the present work we have made use of polarization phase shifting in digital holographic microscopy (DHM) for three dimensional phase profiling of transmissive and reflecting microscopic samples. The Mach–Zehnder arrangement with proper polarizing elements (polarizer-masked cube beam splitter, quarter wave plate and a linear polarizer) is used for recording the phase-shifted digital holograms. The suggested procedure is simple and accurate and obviates the need of piezo devices for phase shifting. The phase profile of the specimen is reconstructed from the holograms by using standard phase shifting algorithms.     

Wave front reconstruction of Fresnel off-axis holograms with compensation of aberrations by means of phase-shifting digital holography

Off-axis holograms recorded with a CCD camera are numerically reconstructed in amplitude by calculating through the Fresnel–Kirchhoff integral. A phase-shifting Mach–Zehnder interferometer is used for recording four-quadrature phase-shifted off-axis holograms. The basic principle of this technique and its experimental verification are described. We show that the application of this algorithm allows for the suppression of the zero order of diffraction and of the twin image and that the contrast of the reconstructed images can be further enhanced by digital compensation of the aberrations introduced by the holographic recording system     

Wave optics analysis by phase-shifting real-time holographic interferometry

The purpose of this work is to study the potentialities in the phase-shifting real-time holographic interferometry using photorefractive crystals as the recording medium for wave-optics analysis in optical elements and non-linear optical materials. This technique was used for obtaining quantitative measurements from the phase distributions of the wave front of lens and lens systems along the propagation direction with in situ visualization, monitoring and analysis in real time.     

Reconstruction algorithms applied to in-line Gabor digital holographic microscopy

This paper investigates the application of Fresnel based numerical algorithms for the reconstruction of Gabor in-line holograms. We focus on the two most widely used Fresnel approximation algorithms, the direct method and the angular spectrum method. Both algorithms involve calculating a Fresnel integral, but they accomplish it in fundamentally different ways. The algorithms perform differently for different physical parameters such as distance, CCD pixel size, and so on. We investigate the constraints for the algorithms when applied to in-line Gabor digital holographic microscopy. We show why the algorithms fail in some instances and how to alter them in order to obtain useful images of the microscopic specimen. We verify the altered algorithms using an optically captured digital hologram.     

Error evaluation for Zernike polynomials fitting based phase compensation of digital holographic microscopy

Combining the experimental research with the simulation calculation, the error evaluation for Zernike polynomials fitting (ZPF) based phase compensation of digital holographic microscopy (DHM) is performed. The obtained results show that the reconstructed phase with high precision can be obtained by ZPF phase compensation algorithm. Moreover, the phase error for ZPF based phase compensation algorithm increases with both the variation of object height and object transverse area, the larger variation of object height, the larger of phase error, and the larger of object transverse area, the faster increase of RMS phase error. To decrease the error of ZPF phase compensation algorithm, it is required to ensure one of the variations of object height and object transverse area to be a small value. Importantly, the proposed method supplies a useful tool for the error evaluation of phase compensation algorithm.     

Optical diffraction tomography microscopy with transport of intensity equation using a light-emitting diode array

Optical diffraction tomography (ODT) is an effective label-free technique for quantitatively refractive index imaging, which enables long-term monitoring of the internal three-dimensional (3D) structures and molecular composition of biological cells with minimal perturbation. However, existing optical tomographic methods generally rely on interferometric configuration for phase measurement and sophisticated mechanical systems for sample rotation or beam scanning. Thereby, the measurement is suspect to phase error coming from the coherent speckle, environmental vibrations, and mechanical error during data acquisition process. To overcome these limitations, we present a new ODT technique based on non-interferometric phase retrieval and programmable illumination emitting from a light-emitting diode (LED) array. The experimental system is built based on a traditional bright field microscope, with the light source replaced by a programmable LED array, which provides angle-variable quasi-monochromatic illumination with an angular coverage of ±37 degrees in both x and y directions (corresponding to an illumination numerical aperture of ∼0.6). Transport of intensity equation (TIE) is utilized to recover the phase at different illumination angles, and the refractive index distribution is reconstructed based on the ODT framework under first Rytov approximation. The missing-cone problem in ODT is addressed by using the iterative non-negative constraint algorithm, and the misalignment of the LED array is further numerically corrected to improve the accuracy of refractive index quantification. Experiments on polystyrene beads and thick biological specimens show that the proposed approach allows accurate refractive index reconstruction while greatly reduced the system complexity and environmental sensitivity compared to conventional interferometric ODT approaches.     

基于相移技术的显微数字全息重构细胞相位

介绍了用显微镜物镜、压电陶瓷和CCD建立的一套测量细胞相位的显微数字全息光路,基于相移技术,给出了重构相位的理论分析,并用洋葱磷片叶细胞作为测试样品,完成了测量细胞相位的实验.结果表明：该系统可以完成细胞相位重构,系统分辨率不低于1μm.     

4-D imaging of fluid flow with digital in-line holographic microscopy

The large depth of field of digital in-line holographic microscopy (DIHM) with numerical reconstruction provides an ideal tool for the study of microfluidic phenomena. As indicators of the flow patterns we use latex microspheres and also red blood cells whose three-dimensional trajectories and velocities can easily be measured as a function of time with subsecond and micron resolution. We demonstrate the efficiency of DIHM by showing 3-D views of the flow patterns around big spheres in various geometric arrangements.     

Design of the spatial filter window for digital holographic convolution reconstruction of object beam field

In the convolution reconstruction process of digital holographic object beam field, the object beam fields with different magnifications can be obtained when the reconstructing beams are spherical waves with different wave curvature radii. This paper presents a theoretical and experimental discussion on the useful spatial frequency spectrum of the hologram numerically illuminated by the spherical wave. The result shows that there would be an image in the spatial frequency spectrum of digital hologram, which is completely the same as the object, if the wave surface radius of the spherical wave is equal to the distance from the object to the CCD sensor. Taking this image as a reference and designing a filter, the position of the reconstructed images with different magnifications can be predicted correctly in the reconstructed plane. Additionally, since the spectrum distribution of zero-order diffraction can be forecasted accurately in theory, its contribution can be effectively eliminated through changing the spatial filter shape; then a reconstructed object field containing more high-frequency information can be obtained.     

Measuring and calculating geometrical parameters of marine plankton using digital laser holographic imaging

The purpose of this paper is to develop an approach using digital laser holographic imaging for measuring and calculating geometrical parameters of marine plankton. It is very important for study of marine plankton and it facilitates the analysis of marine plankton. Using this approach, we obtained geometrical parameters of marine plankton including perimeter, equivalent diameter, lateral size and so on. It is found that a measurement aberration is less than 8% for copepods sample by using collimating reference wave.     

Common-path phase-shifting digital holographic microscopy: A way to quantitative phase imaging and superresolution

We present an experimental setup useful for complex amplitude evaluation and phase image quantification of three-dimensional (3-D) samples in digital holographic microscopy (DHM). It is based on a common-path interferometric configuration performed by dividing the input plane in two contiguous regions and by placing a translation grating near to the Fourier plane. Then, complex amplitude distribution of the sample under test is recovered with phase-shifting standard method obtained by moving the grating using a linear motion stage. Some experimental results of an USAF resolution test are presented for different numerical aperture (NA) microscope lenses. In a second part, the proposed setup is tested under superresolution purposes. Based on the object’s spectrum shift produced by off-axis illumination, we use time multiplexing to generate a synthetic aperture enlargement that improves the final image resolution. Experimental results for the case of a biosample (human red blood cells) and a commercial low NA microscope lens validates the suggested superresolution approach.     

Objective evaluation of images reconstructed from digital in-line Fresnel holograms by using coherent background elimination

Quality of images reconstructed from in-line Fresnel holograms using elimination of a coherent background is quantitatively studied through computer simulation and experiment. The coherent background is digitally calculated by averaging an intensity of a recorded hologram. The results show that in spite of the fact that virtual image is still intact, the background elimination can resolve images with quality comparable to the phase-shifting technique.     

Complex-amplitude-based phase unwrapping method for digital holographic microscopy

A complex-amplitude-based phase unwrapping approach for digital holographic microscopy is proposed in this paper. A quality map is derived directly from the reconstructed complex amplitude distribution of object wave to evaluate noise influence and phase reliability in the wrapped phase image. Quality-guided phase unwrapping algorithm is then implemented with the quality map to retrieve continuous phase profile. Unwrapping errors caused by unreliable phase data are successfully suppressed. The effectiveness of this method is demonstrated with simulation and experimental results.     

Design of adaptive spatial filter at uniform standard for automatic analysis of digital holographic microscopy

The spatial filtering method is a useful technique for the dynamic and automatic analysis in digital holographic microscopy. It has been shown that the proper selection of the spatial filter would improve the quality of the reconstructed image. However different results would be obtained by employing the spatial filter with different parameter threshold decided at different standard. This paper, according to the histogram analysis of the distribution of the +1 term spectrum of each hologram, gropes for the uniform standard for the decision of the threshold of the adaptive filter. It helps the adaptive spatial filtering method to be more advantageous for the dynamic and automatic analysis.     

Method for eliminating zero-order diffraction in lensless Fourier transform digital holography

A method to induce phase shifting in lensless Fourier of digital holography system is presented. In this method, by computer simulation and theoretical analysis on the technology to eliminate the influence of zero-order diffraction in force, it can be found that, a reference light induced in a random non-2π integral number of phase shifting shooting to get the second hologram, and wave reconstruction can be got by the difference value image of the two holograms. And, the method of different digital holography record system with different phase shifting should be used. In this paper, theoretical analyses have been done in detail to discuss the problems that exist in the unsuitable phase-shifting methods. Furthermore, some experiments have been done to prove the reliability of this method. This method can significantly improve the image quality and give better resolution of the reconstructed image.     

Volume-grating phase-shifting digital speckle pattern interferometry used for measurement of out-of-plane displacement field

A new digital speckle pattern interferometry, called volume-grating phase-shifting digital speckle pattern interferometry, is discussed in this paper. The out-of-plane displacement field of a bent plate can be quantitatively measured using volume-grating phase-shifting digital speckle pattern interferometry proposed in this paper. Theoretical and experimental results, as well as absolute errors, are given.     

Correction of quadratic phase error in two-wavelength digital holographic interferometry using laser diodes

We present the correction of a quadratic phase error in two-wavelength digital holographic interferometry using laser diodes. This phase error arises from numerical reconstructions of wavefronts from digital holograms based on the Fresnel diffraction integral. To correct the quadratic phase error, it is numerically produced by computer on the basis of the theoretical prediction and is subtracted from the phase difference map in two-wavelength digital holographic interferometry. Experimental results show that the method of correction in this paper is useful for two-wavelength digital holographic interferometry using laser diodes.     

Reduction of speckle noise in digital holography by using digital image processing

A fundamental problem in optical and digital holography is the presence of speckle noise in the reconstruction process. Many approaches have been carried out in order to overcome such a problem ranging from modifying the spatial coherence of the illumination (optical techniques) to image processing techniques (digital techniques). This work shows the merged use of digital image processing techniques in order to reduce the speckle noise in digital reconstruction of optically recorded Fresnel's holograms. The proposed filtering techniques are illustrated with experimental results.