Three-dimensional microscopy with phase-shifting digital holography (original) (raw)
Space-bandwidth conditions for efficient phase-shifting digital holographic microscopy
Journal of the Optical Society of America A, 2008
Microscopy by holographic means is attractive because it permits true three-dimensional (3D) visualization and 3D display of the objects. We investigate the necessary condition on the object size and spatial bandwidth for complete 3D microscopic imaging with phase-shifting digital holography with various common arrangements. The cases for which a Fresnel holographic arrangement is sufficient and those for which object magnification is necessary are defined. Limitations set by digital sensors are analyzed in the Wigner domain. The trade-offs between the various holographic arrangements in terms of conditions on the object size and bandwidth, recording conditions required for complete representation, and complexity are discussed.
Common-path, single-shot phase-shifting digital holographic microscopy using a Ronchi ruling
Applied Physics Letters, 2019
Phase-shifting digital holography is widely considered to be a groundbreaking method to quantitatively investigate the phase distribution of specimens, such as living cells. The main flaws of this method, however, are that the requirement for several sequential phase-shifted holograms eliminates the possibility of single-shot imaging and complex configurations would also increase the temporal noise. The present paper aims to validate a single-shot, common-path, phase-shifting digital holographic microscopy, employing a self-referencing geometry. A Ronchi ruling, located in the Fourier plane of a standard microscopic imaging system, produces multiple replicas of sample information in the image plane. The phase retrieval algorithm is performed by superposition of the sample-free portion of each replica with the object information, and requires at least three adjacent diffraction orders. To evaluate the performance of the proposed method, the phase distribution of silica microspheres a...
Digital Holographic Microscopy
Optik & Photonik, 2007
Digital holographic microscopy provides quantitative phase contrast imaging that is suitable for high resolving investigations on reflective surfaces as well as for marker-free analysis of living cells. Results from engineered surfaces and living cells demonstrate applications of digital holographic microscopy for technical inspection and life cell imaging.
Digital Holographic Microscopy (DHM)
Imaging & Microscopy, 2006
Digital Holographic Microscopes (DHM) enables strictly noninvasive visualisation of unstained transparent and partially reflective specimens, in real time, by providing simultaneously amplitude and phase changes of a light wave transmitted or reflected. They are used for characterisation of samples at the nanometer scale, for quality control on production line, and for dynamical analysis of biological specimen and micro systems. more than 15 reconstructions per second for 512 x 512 pixels holograms with a standard personal computer.
Incoherent Digital Holographic Microscopy with Coherent and Incoherent Light
Holography is an attractive imaging technique as it offers the ability to view a complete three-dimensional volume from one image. However, holography is not widely applied to the regime of fluorescence microscopy, because fluorescent light is incoherent and creating holograms requires a coherent interferometer system. We review two methods of generating digital Fresnel holograms of three-dimensional microscopic specimens illuminated by incoherent light. In the first method, a scanning hologram is generated by a unique scanning system in which Fresnel zone plates (FZP) are created by a coherently illuminated interferometer. In each scanning period, the system produces an on-axis Fresnel hologram. The twin image problem is solved by a linear combination of at least three holograms taken with three FZPs with different phase values. The second hologram reviewed here is the Fresnel incoherent correlation hologram. In this motionless holographic technique, light is reflected from the 3-D specimen, propagates through a spatial light modulator (SLM), and is recorded by a digital camera. Three holograms are recorded sequentially, each for a different phase factor of the SLM function. The three holograms are superposed in the computer, such that the result is a complex-valued Fresnel hologram that does not contain a twin image. When these two types of hologram are reconstructed in the computer, the 3-D properties of the specimen are revealed.
Applied Optics, 1999
We present a digital method for holographic microscopy involving a CCD camera as a recording device. Off-axis holograms recorded with a magnified image of microscopic objects are numerically reconstructed in amplitude and phase by calculation of scalar diffraction in the Fresnel approximation. For phasecontrast imaging the reconstruction method involves the computation of a digital replica of the reference wave. A digital method for the correction of the phase aberrations is presented. We present a detailed description of the reconstruction procedure and show that the transverse resolution is equal to the diffraction limit of the imaging system.
Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing Xv, 2008
Different interferometric techniques were developed last decade to obtain full field, quantitative, and absolute phase imaging, such as phase-shifting, Fourier phase microscopy, Hilbert phase microscopy or digital holographic microscopy (DHM). Although, these techniques are very similar, DHM combines several advantages. In contrast, to phase shifting, DHM is indeed capable of single-shot hologram recording allowing a real-time absolute phase imaging. On the other hand, unlike to Fourier phase or Hilbert phase microscopy, DHM does not require to record in focus images of the specimen on the digital detector (CCD or CMOS camera), because a numerical focalization adjustment can be performed by a numerical wavefront propagation. Consequently, the depth of view of high NA microscope objectives is numerically extended. For example, two different biological cells, floating at different depths in a liquid, can be focalized numerically from the same digital hologram. Moreover, the numerical propagation associated to digital optics and automatic fitting procedures, permits vibrations insensitive fullfield phase imaging and the complete compensation for a priori any image distortion or/and phase aberrations introduced for example by imperfections of holders or perfusion chamber. Examples of real-time full field phase images of biological cells have been demonstrated.
Imaging in digital holographic microscopy
Optics Express, 2007
We present a theoretical formalism for three dimensional (3D) imaging properties of digital holographic microscopy (DHM). Through frequency analysis and visualization of its 3D optical transfer function, an assessment of the imaging behavior of DHM is given. The results are compared with those from other types of interference microscopy. Digital holographic microscopy does not result in true 3D imaging. The main advantage of holographic microscopy lies in its quick acquisition of a single 2D image. Full 3D imaging can be obtained with DHM using a broad-band source or tomographic reconstruction.
Methods of Fourier optics in digital holographic microscopy
Optics Communications, 2008
In this paper, processing methods of Fourier optics implemented in a digital holographic microscopy system are presented. The proposed methodology is based on the possibility of the digital holography in carrying out the whole reconstruction of the recorded wave front and consequently, the determination of the phase and intensity distribution in any arbitrary plane located between the object and the recording plane. In this way, in digital holographic microscopy the field produced by the objective lens can be reconstructed along its propagation, allowing the reconstruction of the back focal plane of the lens, so that the complex amplitudes of the Fraunhofer diffraction, or equivalently the Fourier transform, of the light distribution across the object can be known. The manipulation of Fourier transform plane makes possible the design of digital methods of optical processing and image analysis. The proposed method has a great practical utility and represents a powerful tool in image analysis and data processing. The theoretical aspects of the method are presented, and its validity has been demonstrated using computer generated holograms and images simulations of microscopic objects.