0000000001305043
AUTHOR
Jorge Garcia-sucerquia
Physical compensation of phase curvature in digital holographic microscopy by use of programmable liquid lens.
Quantitative phase measurements obtained with digital holographic microscopes are strongly dependent on the optical arrangement of the imaging system. The nontelecentric operation provides phase measurements affected by a parabolic phase factor and requires numerical postprocessing, which does not always remove all the perturbation. Accurate phase measurements are achieved by using the imaging system in telecentric mode. Unfortunately, this condition is not accomplished when a commercial microscope is used as the imaging system. In this paper, we present an approach for obtaining accurate phase measurements in nontelecentric imaging systems without the need for numerical postprocessing. The…
Enhancing spatial resolution in digital holographic microscopy by biprism structured illumination.
A novel and efficient architecture of a structured-illumination digital holographic microscope (DHM) is presented. As the DHM operates at the diffraction limit, its spatial resolution on label-free imaging of transparent samples is improved by illuminating the sample with a structured illumination produced by a Fresnel's biprism. The theoretical analysis of the method forecasts a twofold improvement of the spatial resolution. The proposed method requires only two images to improve the spatial resolution, which eases the process of unmixing the high-resolution components by means of an unknown phase-shift procedure. Numerical modeling and experimental results validate the theoretical finding…
Phase-shifting by means of an electronically tunable lens: quantitative phase imaging of biological specimens with digital holographic microscopy
The use of an electronically tunable lens (ETL) to produce controlled phase shifts in interferometric arrangements is shown. The performance of the ETL as a phase-shifting device is experimentally validated in phase-shifting digital holographic microscopy. Quantitative phase maps of a section of the thorax of a Drosophila melanogaster fly and of human red blood cells have been obtained using our proposal. The experimental results validate the possibility of using the ETL as a reliable phase-shifter device.
Femtosecond digital lensless holographic microscopy to image biological samples
The use of femtosecond laser radiation in digital lensless holographic microscopy (DLHM) to image biological samples is presented. A mode-locked Ti:Sa laser that emits ultrashort pulses of 12 fs intensity FWHM, with 800 nm mean wavelength, at 75 MHz repetition rate is used as a light source. For comparison purposes, the light from a light-emitting diode is also used. A section of the head of a drosophila melanogaster fly is studied with both light sources. The experimental results show very different effects of the pinhole size on the spatial resolution with DLHM. Unaware phenomena on the field of the DLHM are analyzed.
Handheld and cost-effective Fourier lightfield microscope
In this work, the design, building, and testing of the most portable, easy-to-build, robust, handheld, and cost-effective Fourier Lightfield Microscope (FLMic) to date is reported. The FLMic is built by means of a surveillance camera lens and additional off-the-shelf optical elements, resulting in a cost-effective FLMic exhibiting all the regular sought features in lightfield microscopy, such as refocusing and gathering 3D information of samples by means of a single-shot approach. The proposed FLMic features reduced dimensions and light weight, which, combined with its low cost, turn the presented FLMic into a strong candidate for in-field application where 3D imaging capabilities are pursu…
Digital holographic microscopy as a screening technology for diabetes
Label-free quantitative phase imaging (QPI) is the hallmark of digital holographic microscopy (DHM). One of the most interesting medical applications of QPI-DHM is that it can be used to analyze illnesses in which the refractive index or/and the morphology of cells/tissues are distorted, from the acquisition of a single image. In this contribution, we obtain the phase maps of red blood cells (RBCs) samples of patients suffering from diabetes mellitus type 1 (DM1) by using a DHM. Our experimental results show that the measured phase values are significantly different between control non-diabetic and diabetic patients. The high correlation coefficient between the phase and the glycated hemogl…
Aberration compensation for objective phase curvature in phase holographic microscopy: comment
In a recent Letter by Seo et al. [Opt. Lett. 37, 4976 (2012)], the numerical correction of the quadratic phase distortion introduced by the microscope objective in digital holographic microscopy (DHM) has been presented. In this comment, we would like to draw to the attention of the authors and the readers in general that this approach could not be the optimal solution for maintaining the accuracy of the quantitative phase via DHM. We recall that the use of telecentric imaging systems in DHM simplifies the numerical processing of the phase images and produces more accurate measurements.
Fast and robust phase-shift estimation in two-dimensional structured illumination microscopy.
A method of determining unknown phase-shifts between elementary images in two-dimensional Structured Illumination Microscopy (2D-SIM) is presented. The proposed method is based on the comparison of the peak intensity of spectral components. These components correspond to the inherent structured illumination spectral content and the residual compo- nent that appears from wrongly estimated phase-shifts. The estimation of the phase-shifts is carried out by finding the absolute maximum of a function defined as the normalized peak intensity difference in the Fourier domain. This task is performed by an optimization method providing a fast estimation of the phase-shift. The algorithm stability an…
Resolution limit in opto-digital systems revisited
The resolution limit achievable with an optical system is a fundamental piece of information when characterizing its performance, mainly in case of microscopy imaging. Usually this information is given in the form of a distance, often expressed in microns, or in the form of a cutoff spatial frequency, often expressed in line pairs per mm. In modern imaging systems, where the final image is collected by pixelated digital cameras, the resolution limit is determined by the performance of both, the optical systems and the digital sensor. Usually, one of these factors is considered to be prevalent over the other for estimating the spatial resolution, leading to the global performance of the imag…
Recent Advances in Digital Holographic Microscopy
In digital holographic microscopy (DHM) a hologram is captured in the image space provided by a microscope. The transfer of the phase and amplitude structure in the original sample to the hologram is in fact strongly affected by the use of the imaging microscope. A big research effort has been devoted to correct these distortions both by numerical and optical compensation. In this contribution, we present several proposals to improve the performance of classical DHM architectures by an a priori approach to compensate physically these perturbations. Experimental results are also presented to validate the proposed techniques.
View images with unprecedented resolution in integral microscopy
Integral microscopy is a novel technique that allows the simultaneous capture of multiple perspective images of microscopic samples. This feature is achieved at the cost of a significant reduction of the spatial resolution. In fact, it is assumed that in the best cases the resolution is reduced by a factor that is not smaller than ten, what poses a hard drawback to the utility of the technique. However, to the best of our knowledge, this resolution limitation has never been researched rigorously. For this reason, the aim of this paper is to explore the real limitations in resolution of integral microscopy and to obtain optically, without the need of any image-processing algorithm, perspecti…
Off-axis digital holographic microscopy: practical design parameters for operating at diffraction limit.
The utilization of microscope objectives (MOs) in digital holographic microscopy (DHM) has associated effects that are not present in conventional optical microscopy. The remaining phase curvature, which can ruin the quantitative phase imaging, is the most evident and analyzed. As phase imaging is considered, this interest has made possible the development of different methods of overcoming its undesired consequences. Additionally to the effects in phase imaging, there exist a set of less obvious conditions that have to be accounted for as MOs are utilized in DHM to achieve diffraction-limit operation. These conditions have to be considered even in the case in which only amplitude or intens…
Diabetes screening by telecentric digital holographic microscopy
Diabetes is currently the world's fastest growing chronic disease and it is caused by deficient production of insulin by the endocrine pancreas or by abnormal insulin action in peripheral tissues. This results in persistent hyperglycaemia that over time may produce chronic diabetic complications. Determination of glycated haemoglobin level is currently the gold standard method to evaluate and control sustained hyperglycaemia in diabetic people. This measurement is currently made by high-performance liquid chromatography, which is a complex chemical process that requires the extraction of blood from the antecubital vein. To reduce the complexity of that measurement, we propose a fully-optica…
Shift-variant digital holographic microscopy: inaccuracies in quantitative phase imaging
Inaccuracies introduced in quantitative phase digital holographic microscopy by the use of nontelecentric imaging systems are analyzed. Computer modeling of the experimental result shows that even negligible errors in the radius and center of curvature of the numerical compensation needed to get rid of the remaining quadratic phase factor introduce errors in the phase measurements; these errors depend on the position of the object in the field-of-view. However, when a telecentric imaging system is utilized for the recording of the holograms, the numerical modeling and experimental results show the shift-invariant behavior of the quantitative-phase digital holographic microscope.
Optical sectioning with a Wiener-like filter in Fourier integral imaging microscopy
Non-scanning, single-shot, 3D integral microscopy with optical sectioning is presented. The method is based on the combination of Fourier-mode integral microscopy with a 3D deconvolution technique. Specifically, the refocused volume provided by a regular back-projection algorithm is 3D deconvolved with a synthetic 3D impulse response function that takes into account the number and positions of the elemental images. The use of this hybrid technique provides a stack of true-color depth-refocused images with significant gain of optical sectioning. The stack can be used, among other applications, to inspect inside the thick microscope specimen, to calculate collections of perspective views wit…
FIMic: design for ultimate 3D-integral microscopy of in-vivo biological samples
In this work, Fourier integral microscope (FIMic), an ultimate design of 3D-integral microscopy, is presented. By placing a multiplexing microlens array at the aperture stop of the microscope objective of the host microscope, FIMic shows extended depth of field and enhanced lateral resolution in comparison with regular integral microscopy. As FIMic directly produces a set of orthographic views of the 3D-micrometer-sized sample, it is suitable for real-time imaging. Following regular integral-imaging reconstruction algorithms, a 2.75-fold enhanced depth of field and [Formula: see text]-time better spatial resolution in comparison with conventional integral microscopy is reported. Our claims …
3D Integral Microscopy based in far-field detection
Lately, Integral-Imaging systems have shown very promising capabilities of capturing the 3D structure of micro- scopic and macroscopic scenes. The aim of this work is to provide an optimal design for 3D-integral microscopy with extended depth of field and enhanced lateral resolution. By placing an array of microlenses at the aperture stop of the objective, this setup provides a set of orthographic views of the 3D sample. Adopting well known integral imaging reconstruction algorithms it can be shown that the depth of field as well as spatial resolution are improved with respect to conventional integral microscopy imaging. Our claims are supported on theoretical basis and experimental images …
Accurate single-shot quantitative phase imaging of biological specimens with telecentric digital holographic microscopy
The advantages of using a telecentric imaging system in digital holographic microscopy (DHM) to study biological specimens are highlighted. To this end, the performances of nontelecentric DHM and telecentric DHM are evaluated from the quantitative phase imaging (QPI) point of view. The evaluated stability of the microscope allows single-shot QPI in DHM by using telecentric imaging systems. Quantitative phase maps of a section of the head of the drosophila melanogaster fly and of red blood cells are obtained via single-shot DHM with no numerical postprocessing. With these maps we show that the use of telecentric DHM provides larger field of view for a given magnification and permits more acc…
Study of spatial lateral resolution in off-axis digital holographic microscopy
The lateral resolution in digital holographic microscopy (DHM) has been widely studied in terms of both recording and reconstruction parameters. Although it is understood that once the digital hologram is recorded the physical resolution is fixed according to the diffraction theory and the pixel density, still some researches link the resolution of the reconstructed wavefield with the recording distance as well as with the zero-padding technique. Aiming to help avoiding these misconceptions, in this paper we analyze the lateral resolution of DHM through the variation of those two parameters. To support our outcomes, we have designed numerical simulations and experimental verifications. Both…
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