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3D analysis of defects in integrated circuits by one-photon optical-beam-induced current imaging and laser confocal reflectance microscopy
Jelda Jayne Miranda,
Caesar Saloma
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Three-dimensional analysis is performed on defects found in an integrated circuit from high-contrast images that are obtained via an inexpensive technique that combines confocal reflectance microscopy with one-photon optical beam-induced current (1P-OBIC) imaging. The same focused beam simultaneously produces the 1P-OBIC and reflectance signals from the illuminated spot. Exclusive 3D distributions of the metal and semiconductor sites in the vicinity of defects caused by electrical overstress (decrease in OBIC current) and unwanted formation of generation centers (increase in OBIC current), reveal features which are difficult to isolate with confocal or 1P-OBIC microscopy alone.
The use of advanced light source and signal processing in scanning optical microscopy
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Ultrafast lasers have found increasing use in scanning optical microscopy because of its very high peak power in generating multiphoton optical excitations. We are demonstrating that the multiphoton processes can be further extended to third harmonic generation (THG) and two-photon (2-p) excitation in the UVB range with broader tunability enabled by a synchronously pumped optical parametric oscillator (OPO). The scanning nature of image acquisition process also greatly facilitates the incorporation of techniques in signal processing, which opens further possibilities. For example, the very short pulse width of ultrafast lasers allows excitation and sampling of photo-processes with extremely broad bandwidth. As such, radio frequency is excited as a contrast signal in imaging high speed photodiodes. In addition, dithering techniques that base on lock-in detection allows signal conditioning so that better signal to noise ratio can be resulted and features of high spatial frequency can be emphasized.
Probe dendritic functions through poking and peeking
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Several photonic approaches have been utilized to study functional dynamics of olfactory bulb dendrites, which plays a critical role in odor discrimination and recognition. Firstly, with infrared differential interference contrast (DIC) video microscopy, we can visualize living nerve cells in an olfactory bulb slice preparation and target glass electrodes to different dendritic locations for direct electrical measurement. This furnishes a high temporal resolution of signal recording from dendrites. Secondly, by using a cooled CCD camera and loading calcium-sensitive dyes into neurons, we have explored the spatial distribution and propagation of spike signals within complex dendritic trees. Thirdly, two-photon microscope enables us to analyze active properties of very tiny dendritic structures such as dendritic spines. Lastly, by using UV light pulse to release calcium ions from caged compounds, we have examined the mechanisms for signal communication between two dendrites with reciprocal synaptic connections. Our research highlights an important contribution of optical imaging methods to functional dissection of neuronal circuitry in the brain.
Application of confocal microscopy in neurons imaging
Xiaoxiang Zheng
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Nitric oxide (NO) and calcium ions (Ca2+) play critical role as molecular mediator in many physiological processes. However, their low concentration and instability in specimen make them difficult to be detected directly in neurons. We developed a method for imaging nitric oxide and calcium ions using Laser Scanning Confocal Microscopy (LSCM). Cultured hippocampal neuron is dyed and observed under Zeiss LSM 510 laser scanning confocal microscope. Excited by laser the emission light from the labeled nitric oxide and calcium ions in the neutron are detected. In this way, the nitric oxide and calcium ions are imaged and their intracellular kinetic change in monitored. Furthermore, image processing and visualization techniques are employed to help analyze the image data.
Particle-fixed Monte Carlo model for optical coherence tomography
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Optical Coherence Tomography is a new technique mainly used in biomedical imaging. Here we present a Particle-Fixed Monte Carlo (PFMC) simulation for OCT signal. In PFMC model the scattering particles of the sample are assumed to be temporarily fixed randomly in simulation process of the backscattering light. The new model, beyond the convention Monte Carlo simulation, explains very well the exponential decay signal at the interface of different media layers in OCT experimental measurement.
Automatic detection of intracellular nitric oxide concentration by confocal laser scanning microscopy images analysis
John Kutor,
Gang-min Ning,
Di-hui Hong,
et al.
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Confocal laser scanning microscopy (CLSM) is one of the common equipments being used to observe the intracellular substances like nitric oxide (NO) and calcium ions (Ca2+). The intracellular substances detection relies on the proper analysis of the fluorescence images obtained by CLSM. At present, the outlines of the cells in CLSM images from fluorescence are manually drawn by judging the contrast between the object and the background. This method is subjective and may have the possibility of inaccuracy. In this paper we have designed a technique based on digital imaging processing to automatically detect the outlines or contours of the cells and assess the production of NO labeled by fluorescence probes. The developed technique was tested on a series of images obtained from CLSM and the spread of NO during the time course was satisfactorily estimated.
Improvement of the resolution and instantaneous fringe-order identification of optical coherence tomography by optimum wavelength combination
Zhongbao Xu,
Hunian Ye,
Jianping Song
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We report a novel technique in which by optimum wavelength combination from dual broadband optical sources obtains a short coherence length for use in optical coherence tomography (OCT) systems. Another advantages of this technique lie in the ability to identify correctly the center fringe position, as a result, such as position, displacement or an absolute phase measurement over a large operating can be achieved. Theoretical analysis, computer simulations, and experimental verifications have shown that these techniques are able to increase greatly the dynamic range of the measurement under a low signal-to-noise ratio environment.
Non-invasive optical imaging of tumor growth in intact animals
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We describe here a system for rapidly visualizing tumor growth in intact rodent mice that is simple, rapid, and eminently accessible and repeatable. We have established new rodent tumor cell line -- SP2/0-GFP cells that stably express high level of green fluorescent protein (GFP) by transfected with a plasmid that encoded GFP using electroporation and selected with G418 for 3 weeks. 1 x 104 - 1x107 SP2/0-GFP mouse melanoma cells were injected s.c. in the ears and legs of 6- to 7-week-old syngeneic male BALB/c mice, and optical images visualized real-time the engrafted tumor growth. The tumor burden was monitored over time by cryogenically cooled charge coupled device (CCD) camera focused through a stereo microscope. The results show that the fluorescence intensity of GFP-expressing tumor is comparably with the tumor growth and/or depress. This in vivo optical imaging based on GFP is sensitive, external, and noninvasive. It affords continuous visual monitoring of malignant growth within intact animals, and may comprise an ideal tool for evaluating antineoplastic therapies.
Tussue birefringence characterization by use of fiber-based polarization-sensitive optical coherence tomography
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A fiber-based polarization-sensitive optical coherence tomography system was described. The polarization modulator in this system was introduced in the reference arm rather than in the source arm, providing an increased power delivered from light source to sample. Based on angle preservation of Stokes vectors in the PS-OCT system, Stokes parameters of backscattered light measured at the detection arm were used to determine the phase retardation of birefringence samples as a function of depth. With the developed PS-OCT system, investigation on birefringence alternation of ligament under different physical condition was carried out.
Fluorescence correlation spectroscopy based upon two-photon excitation
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Fluorescence correlation spectroscopy (FCS) is a powerful tool for measurement of biological dynamic processes. In this studying, a two-photon excitation fluorescence correlation spectroscopy (TP-FCS) system was set up depending on a part of optical block and detector of the multi-photon excitation fluorescence microscope (MPLFM). The phenomenon "photon-burst" was observed from the TP-FCS system. Meanwhile, the diffusion coefficient of rhodamine B molecule in sucrose aqueous solution was calculated. It was proved that TP-FCS is especially suited for integration into MPEFM accordingly to yield a hybride-technology with the peculiarities of the individual technique and the advantage of mutual synergistic effects. The fusion of both techniques seems to be reasonable and desirable to reduce costs.
The fluorescent photobleaching properties of GFP expressed in human lung cancer cells
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The characteristic properties of GFP make this protein a good candidate for use as a molecular reporter to monitor patterns of protein localization, gene expression, and intracellular protein trafficking in living cells. In this study, the dicistronic expression vector (pEGFP-C1) was used to transfected into human lung cancer cell line (ASTC-a-1) and a positive clone which stably expressed GFP in high level was obtained. After more than three months' passengers, the cells were also remained the strong fluorescence under fluorescent microscope. The results showed that the green fluorescent protein expressed in tumor cells was also photobleached under intense irradiation (approximately 488 nm) and the degree of photobleaching varied with the difference of the intensity of the excitation. Using different interdiction parcel (None, ND4, ND8, ND16), there were significant differences in photobleaching among the different excitation. The photobleaching was also affected by the time length of excitation, and the intensity of fluorescence was obviously decreased along with the increasing of excitation time, especially to stronger excitation.
Tissue elaborate structure data collection by a high-resolution optical method and image reconstruction
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Optical coherence tomography (OCT) uses low coherence interferometric techniques to obtain high resolution reflectivity profiles of the sample without any damnification. According to the "optically sectioning" of the specimen and right way for 3D reconstruction, we can get careful configuration of biological tissues. OCT as a tool for 3D optical microscopy such as X-ray, ultrasound will be an important part of biological tissues imaging studies.
Detection of esophageal cancer by human serum using autofluorescence and Raman spectroscopy
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We detected the serum of patient's and normal using auto-fluorescence and Raman spectroscopy. The serum spectrum was excited by laser of the wavelength 488.0 nm and 514.5 nm. Compared with the normal serum, we can observe the apparent differences of autofluorescence-Raman spectroscopy: the majority of the fluorescence spectrum did not have violent alteration, but three Raman peak had disappeared or very weak. After operation, the Raman spectrum of patient's serum is similar to the normal. And fluorescence peak's red shift, α-value also provide the reference for future research. We assume that following the progression of the tumor, β-value will decrease following the aggravation of cancer. The result of spectrum analysis is accordance with the clinical diagnosis.
Analysis of normal and diseased liver tissue using auto-fluorescence and Raman spectroscopy
Xiaozhou Li,
Chunde Jia,
Junxiu Lin,
et al.
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In this paper, laser induced human serum Raman spectra of liver cancer are measured. The spectra differences in serum from normal people and liver cancer patients are analyzed. For the typical spectrum of normal serum, there are three sharp Raman peaks and relative intensity of Raman peaks excited by 514.5 nm is higher than that excited by 488.0 nm. However, for the Raman spectrum of liver cancer serum there are no peaks or very weak Raman peaks at the same positions. Results from more than two hundred case measurements show that clinical diagnostic accuracy is 92.86%. And then, the liver fibrosis and liver cirrhosis are studied applying the technology of LIF. To liver cirrhosis, the shape of Raman peak is similar to normal and fluorescence spectrum is similar to that of liver cancer from statistic data. The experiment indicates that there is notable fluorescence difference between the abnormal and normal liver tissue and have blue shift in fluorescence peak. These results have important reference values to explore the method of laser spectrum diagnosis.
Detection and characterization of stomach cancer and atrophic gastritis with fluorescence and Raman spectroscopy
Xiaozhou Li,
Junxiu Lin,
Chunde Jia,
et al.
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In this paper, we attempt to find a valid method to distinguish gastric cancer and atrophic gastritis. Auto-fluorescence and Raman spectroscopy of laser induced (514.5 nm and 488.0 nm) was measured. The serum spectrum is different between normal and cancer. Average value of diagnosis parameter for normal serum, red shift is less than 12 nm and Raman relative intensity of peak C by 514.5 nm excited is stronger than that of 488.0 nm. To gastric cancer, its red shift of average is bigger than 12 nm and relative intensity of Raman peak C by 514.5 nm excited is weaker than that by 488.0 nm. To atrophic gastritis, the distribution state of Raman peaks is similar with normal serum and auto-fluorescence spectrum's shape is similar to that of gastric cancer. Its average Raman peak red shift is bigger than 12 nm and the relative intensity of peak C by 514.5 excited is stronger than that of by 488.0. We considered it as a criterion and got an accuracy of 85.6% for diagnosis of gastric cancer compared with the result of clinical diagnosis.
Laser-induced human serum fluorescence and Raman spectroscopy detection of colon cancer
Xiaozhou Li,
Junxiu Lin,
Chunyu Wu,
et al.
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Laser-induced fluorescence and Raman spectroscopy of serum for diagnostic colon cancer was investigated in this paper. The difference to serum spectrum was discovered. Three Raman peaks were consistently observed from normal blood serum emission using 488.0 nm and 514.5 nm excitation of an Ar-ior laser. While no Raman peak or slight Raman peaks were detected from colon cancer's cases. In addition, the red shift of fluorescence peak and decrease of fluorescence intensity are founded after samples radiated by laser. 120 colon cancer cases are investigated in this paper. Through three parameters we obtained an accuracy of 82.5% compared to clinical diagnosis. These results have important reference values to explore the method of Raman spectrum and LIF for diagnosis of cancer.
Medical microscopic image matching based on relativity
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In this paper, an effective medical micro-optical image matching algorithm based on relativity is described. The algorithm includes the following steps: Firstly, selecting a sub-area that has obvious character in one of the two images as standard image; Secondly, finding the right matching position in the other image; Thirdly, applying coordinate transformation to merge the two images together. As a kind of application of image matching in medical micro-optical image, this method overcomes the shortcoming of microscope whose visual field is little and makes it possible to watch a big object or many objects in one view. Simultaneously it implements adaptive selection of standard image, and has a satisfied matching speed and result.
Investigation of Ar+-laser-induced blood fluorescence spectra
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Laser-blood cells interaction was studied by Ar+ laser-induced animal (mouse) blood fluorescence spectra in vitro. The fluorescence spectra of the blood under various irradiated powers of Ar+ laser excitation are shown that there are very rich and sharp spectral peaks from 600 nm to 860 nm. These peaks are located at 616 nm, 666 nm, 708 nm, 739 nm, 752 nm, 766 nm, 800 nm, 812 nm and 844 nm. This may be due to the fact that there are various fluorophores in the blood and the ground electronic state of the fluorophores containing a large number of vibrational levels. In addition, 666 nm peak among them is the most prominent and is a larger change in the intensity under different power of Ar+ lasers excitation. It foretell that laser near a wavelength of 666 nm may be more effective to low lever laser therapy (LLLT). Furthermore, these experiments indicate that when the laser irradiated power density reaches to 30 mW/cm2 the blood cells are still not destroyed. The results may be significative for the choice of irrediation-wavelength in LLLT.
Quantum mechanics analysis of damping effect on laser-biomolecule interaction
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On the basis of study on resonance exciting action of laser with chains and bonds of biomolecules the in reference damping effect of laser with biomolecules will be further analyzed and discussed using quantum mechanics in this paper.
Analysis of nonlinear behavior of laser-DNA molecule interaction
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Nonlinear chaotic behavior of laser-DNA interaction is analysis by means of Menikov method, frequency spectrum analysis method and Lyapunov exponent Numerical method etc. It is very useful on explanation of genetic mutagene The Department of Physics, Mengzi Teacher College, Mengzi 661100, China sis of laser.