A study was carried out focusing on the effect of the background estimation on the performance of the Tikhonov-Miller algorithm (ICTM), Carrington, and Richardson-Lucy algorithms. A comparison was also made with the l...
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ISBN:
(纸本)0819430757
A study was carried out focusing on the effect of the background estimation on the performance of the Tikhonov-Miller algorithm (ICTM), Carrington, and Richardson-Lucy algorithms. A comparison was also made with the linear Tikhonov-Miller restoration filter. It was found that an underestimation of the background will make the non-negativity constraint ineffective which results in a performance that does not differ much from the performance obtained by the linear restoration filter. A novel method was devised to estimate the background based on the dependency of non-linear restoration algorithms on the background.
A well-known distortion of objects in three-dimensionalmicroscopy manifests itself as an elongation in the axial direction. Authors such as Visser and Hell have seemingly contradicted one another on the cause as well...
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ISBN:
(纸本)0819430757
A well-known distortion of objects in three-dimensionalmicroscopy manifests itself as an elongation in the axial direction. Authors such as Visser and Hell have seemingly contradicted one another on the cause as well as the magnitude of the effect. We have examined these theories and performed simulations and experimental measurements to better understand the nature of the effect. We simulate point spread functions (based on the work of Gibson) taking into account the various refractive indices involved as well as the magnification, the numerical aperture, the working distance of the objective, the depth of the object under the coverslip, and the object's size. We measure the axial and lateral dimensions of digitized images of microspheres that have been 'acquired' using a simulated point spread function that changes as the depth of the object changes. These simulations are done for conventional (optical sectioning) microscopy as well as for confocal microscopy. Further, we have performed experimental measurements on real microspheres on a conventional microscope to relate theory, simulation, and practice. Our measurements and simulations show that 1) the object's size, 2) its depth under the coverslip, 3) the refractive index mismatch between the immersion fluid (nimmersion) and embedding material for the object (nembedded), and 4) the NA of the lens play a pivotal role in the effect.
The optical quadrature imaging technique, as derived and extended from microwave and laser radar quadrature detection techniques, provides an efficient method for obtaining phase information from a sample that has lit...
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ISBN:
(纸本)0819430757
The optical quadrature imaging technique, as derived and extended from microwave and laser radar quadrature detection techniques, provides an efficient method for obtaining phase information from a sample that has little or no amplitude contrast. We are able to resolve internal structures of a sample that are defined by relatively small refractive index differences without the use of dyes or stains, while using much lower light levels than conventional techniques. We have constructed a prototype system for imaging microscopic phase-only objects. In this paper, we present its capabilities, as well as the imaging and reconstruction methods used to obtain quantitative information about a sample.
New method of a tomographic microscopy with spatially incoherent illumination is proposed. Earlier, in our works for reconstruction of a refractive index with spatial distribution of 3D phase samples, a method of tomo...
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ISBN:
(纸本)0819430757
New method of a tomographic microscopy with spatially incoherent illumination is proposed. Earlier, in our works for reconstruction of a refractive index with spatial distribution of 3D phase samples, a method of tomographic microscopy with the coherent light was used. Main disadvantage of this scheme is use of a coherent light, which brings to appearance the phase noise in projection data. Spatially incoherent tomographic microscope is based on the Linnik phase-shifting interference microscope reflected type. Using interferometry with large source of the monochromatic light allows greatly improve a quality of phase projections by averaging of interference patterns. For tomographic mode of Linnik microscope working and obtaining the oblique illumination of sample method for displacing a light source is used. Observation angle range for 100×oil-immersion objective, N.A.=1.25, is 90 degrees. This method allows realizing a two-dimensional scanning trajectory for sample observation. Particularities of tomography for the phase objects, placed near plane mirror, are considered. So, a viewing angle range can be reduced to 90 degree, and sample is complement by its mirror reflection. The iterative algorithms for limited-angle tomographic reconstruction were used. Experimental results on three-dimensional reconstruction of the single human blood cell (erythrocyte) are presented.
Recently a novel imaging technique based on third-harmonic generation (THG) was introduced1-3. This technique relies on a third-order non-linear interaction to generate a coherent signal response on the third-harmonic...
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ISBN:
(纸本)0819430757
Recently a novel imaging technique based on third-harmonic generation (THG) was introduced1-3. This technique relies on a third-order non-linear interaction to generate a coherent signal response on the third-harmonic frequency with respect to the fundamental input radiation. Here we report on the input NA dependence of the THG signal and examine the resulting imaging characteristics of this novel technique in terms of resolution and contrast generation. We'll demonstrate the potential of the technique through a number of imaging examples, with special emphasis on in vivo applications. The latter illustrates the non-invasive character of the technique.
An important approach dealing with sub lambda super resolution effect is based on a 3-D scanning of the examined sample using a pipette with small diameter. The scanning in the z direction is required for the conserva...
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ISBN:
(纸本)0819430757
An important approach dealing with sub lambda super resolution effect is based on a 3-D scanning of the examined sample using a pipette with small diameter. The scanning in the z direction is required for the conservation of the distance between the sample and the pipette and it is time consuming. In this paper we suggest a modification of that approach based on 2-D scanning. The scanning in the z direction is replaced by the usage of white light illumination. Based on the spectrum of the scanned information, one can compute the height of the surface. This consumes much less time than the conventional pipette scanner. This approach appears to be reasonable if the profile is varying not more than a portion of a wavelength. If, however, the object is not flat but slightly curved on a large scale we propose to employ a servo for the height. This servo can be much slower than the signal servo in the traditional approach.
The structural elucidation of complex systems may be simplified with multi-dimensional spectroscopic techniques with some combination of spatial and spectral resolution. Raman spectroscopy permits the addition of anot...
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ISBN:
(纸本)0819430757
The structural elucidation of complex systems may be simplified with multi-dimensional spectroscopic techniques with some combination of spatial and spectral resolution. Raman spectroscopy permits the addition of another variable to this scenario -excitation wavelength. Data obtained using excitation wavelengths from the UV (244 nm) to near-IR (785 nm) regions will be presented showing the qualitative and quantitative study of diamond-like carbon (DLC), silicon, and other systems of an industrial or biomedical nature. The choice of appropriate wavelength provides an additional advantage over other spectroscopic techniques for elucidating specific structural information from these systems. The advantages of UV-Raman for materials science and thin film studies will be considered. The design of instruments and probes for the application of Raman spectroscopy to industrial process control and the development of Raman spectroscopic libraries for contaminant analysis will be discussed.
A previous paper by the authors presented an algorithm that successfully segmented organs grown in vitro from their surroundings. It was noticed that one difficulty in standard dyeing techniques for the analysis of co...
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ISBN:
(纸本)0819430757
A previous paper by the authors presented an algorithm that successfully segmented organs grown in vitro from their surroundings. It was noticed that one difficulty in standard dyeing techniques for the analysis of contours in organs was due to the fact that the antigen necessary to bind with the fluorescent dye was not uniform throughout the cell borders. To address these concerns, a new fluorescent technique was utilized. A transgenic mouse line was genetically engineered utilizing the hoxb7/gfp (green fluorescent protein). Whereas the original technique (fixed and blocking) required a numerous number of noise removal filtering and sophisticated segmentation techniques, segmentation on the GFP kidney required only an adaptive binary threshold technique which yielded excellent results without the need for specific noise reduction. This is important for tracking the growth of kidney development through time.
High intensity chirped pulses can be used for probing microscopic chemical environments through the use of a particular choice of dye, for instance SNAFL2. The basis for this technique is that the excited state popula...
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ISBN:
(纸本)0819430757
High intensity chirped pulses can be used for probing microscopic chemical environments through the use of a particular choice of dye, for instance SNAFL2. The basis for this technique is that the excited state populations can be manipulated through control over the temporal order of the excitation frequencies in the excitation pulse - i.e. chirp - with the outcoming fluorescence as the reporting parameter. A chirp dependent fluorescence response can also be observed in larger molecular systems with more degrees of freedom like for instance green fluorescent proteins. In preparation for application of the technique to microscopy we use a facility permitting observation of this phenomenon in various dyes with high sensitivity. High power, 30 fs pulses from an OPA, tunable from 400 nm to 1.5 micron are used. These pulses with a repetition rate of 1 kHz are sufficiently intense that a relatively large sample region can be excited to saturation from which then a sub-region with uniform excitation conditions can be selected for signal collection.
This Volume 3261 of the conference proceedings contains 36 papers. Topics discussed include optical instrumentation, microscopies for challenging biological imaging and measurement, improved methods and instruments fo...
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This Volume 3261 of the conference proceedings contains 36 papers. Topics discussed include optical instrumentation, microscopies for challenging biological imaging and measurement, improved methods and instruments for biological visualization, multidimensionalimage reconstruction and restoration and multispectral imaging.
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