This study aims at exploring the potential of ATR-far-ultraviolet (FUV) spectroscopy in investigating electronic structure and transitions of various kinds of biological molecules. For this purpose, ATR-FUV spectra were measured for several kinds of proteins with the different secondary structures, several kinds of carbohydrates, nucleic acids, and lipids. Band assignments have been made for all kinds of biological molecules investigated based on our previous ATR-FUV studies on n-alkanes, alcohols, esters, and amides. For example, the proteins show a characteristic band near 200 nm due to π-π* transition of amide groups. The position of this band varies a little with the secondary structure of proteins but its intensity changes significantly depending on the secondary structure and solutions. All the carbohydrates studied yielded a band near 170 nm due to n-Rydberg transition of ether. In addition, acetylcarbohydrates give an additional band near 190 nm originating from π-π* transition of amide at 2’ carbon. The present study has demonstrated that ATR-FUV spectroscopy is a new powerful technique in exploring electronic structure and transitions of biological molecules, in general. It is also possible to use ATR-FUV spectroscopy for quantitative and qualitative analysis of biological molecules. Moreover, it is of note that information regarding electronic transitions collected by ATR-FUV spectroscopy is useful for UV resonance Raman (UVRR) spectroscopy studies of biological molecules. A combined ATR-FUV spectroscopy and UVRR spectroscopy method may provide a novel analytical tool for molecular and electronic structure of biological molecules.
Virus infection of a human cell was determined only 3 h after invagination. We used viral vector Ad-CMV-control (AdC), which lacks the E1 gene coding for early polypeptide 1 (E1). AdC can replicate in human embryonic kidney 293 (HEK293) cells into which the E1 gene has been transfected. According to partial least-square regression discriminant analysis, it was assumed that two kinds of reaction take place in the cell during viral invasion. The first response of the cell was determined 3 h after the virus invasion, and the second one was determined ∼9 h later. The first one seems to be due to compositional changes in DNA. Analysis of large-scale datasets strongly indicated that the second reaction can be attributed to a reduction in protein concentration or uptake of phenylalanine into the nucleus.
The purpose of this study is to develop a estimation technique of local cell activity in cultured 3D cell aggregate with gelatin hydrogel microspheres by using Raman spectroscopy. It is an invaluable technique allowing real-time, nondestructive, and invasive measurement. Cells in body generally exist in 3D structure, which physiological cell-cell interaction enhances cell survival and biological functions. Although a 3D cell aggregate is a good model of the cells in living tissues, it was difficult to estimate their physiological conditions because there is no effective technique to make observation of intact cells in the 3D structure. In this study, cell aggregates were formed by MC3T-E1 (pre-osteoblast) cells and gelatin hydrogel microspheres. In appropriate condition MC3T-E1 cells can differentiate into osteoblast. We assume that the activity of the cell would be different according to the location in the aggregate because the cells near the surface of the aggregate have more access to oxygen and nutrient. Raman imaging technique was applied to measure 3D image of the aggregate. The concentration of the hydroxyapatite (HA) is generated by osteoblast was estimated with a strong band at 950-970 cm-1 which assigned to PO43- in HA. It reflects an activity of the specific site in the cell aggregate. The cell density in this specific site was analyzed by multivariate analysis of the 3D Raman image. Hence, the ratio between intensity and cell density in the site represents the cell activity.
Colorectal adenoma (CA) is a disease caused by various factors (such as genetic factors or environmental exposures). The appearance of colon polyp (CP) within colorectal might indicate the hint of CA development. Ball-lens hollow fiber Raman probe (BHRP) may has a high capability for detection of CA in living experimental animal and have already tested to rat’s CP in this study, which was designed to collaborate between BHRP with mini-endoscopy to observe the biochemical alteration within normal colon tissue and rat’s colon polyps in real time. BHRP and mini-endoscopy can distinguish the differences in their finger print spectra and make pictures the control and CP in the real time. At the first step, the real situation of normal colon and Rat’s CP were washed by saline and observed with mini-endoscopy. BHRP was introduced to Dextran sodium sulphate (DSS)-induced Rat's CP to detect some of biochemical alteration. The main purpose of this study was to introduce mini-endoscopy to guide the BHRP for diagnosing of CP in real time and to compare it with spectra of normal colon (control group) in living rat. As the result, BHRP can provide the differences in band of control and CP group, which can inform that the biochemical of normal and CP has changed. As a major parameter to distinct normal and CP tissue were phosphatidylinositol, phosphodiester group, lipid, and collagen. Mini endoscopy and BHRP is very sensitive devices for diagnosing of CP in real time.
The present study demonstrates that Raman spectroscopy is a powerful tool for the detection of virus-infected cells. Adenovirus infection of human embryonic kidney 293 cells was successfully detected at 12, 24, and 48 h after initiating the infection. The score plot of principal component analysis discriminated the spectra of the infected cells from those of the control cells. The viral infection was confirmed by the conventional immunostaining method performed 24 h after the infection. The newly developed method provides a fast and label-free means for the detection of virus-infected cells.
The combination of confocal micro-Raman spectroscopy and multivariate analysis is carried out for analysis of
maturation of neurons. This study suggests that Raman data reflects the stages of neural maturation which relates with
the expression of new neural function such as spontaneous activity. Neurons obtained from a hippocampus of rat
embryos are cultured in a dish with quartz bottom. According to the previous electrophysiological study, matured neural
cell network showed regulated pulsation with interval of several seconds without any stimulation. It suggested that
alterations in the molecular composition took place in the cell. The Raman measurements are carried out to observe this
alteration along with the maturing process of neurons. The spectra of live neural cells measured after 2, 8, 15, 30, 45, 60,
75, 90, 105 and 120 days of culturing are analyzed by principal component analysis (PCA). The result shows several
groups suggesting the maturation scheme which is observed by the electrophysiological studies. It demonstrates that the
maturation process of neural cells can be monitored by Raman spectroscopy.
Sexual reproductive body can be produced from a fertilized ovum. Once the ovum is fertilized with sperm, it runs
through the cell division, differentiates to all kinds of cells, and goes to make a complete body. However, not all of them are
viable and some of them stop to ontogenesis showing the developmental abnormality. In order to discriminate the egg
quality, we apply Raman spectroscopy for fish egg. After the measurement, these Raman data are checked up with the
information about the eggs can survive or not, and we examine what factors are important in egg components to distinguish
between “good quality” and “not good quality”. We present the results of assessment of egg quality, and investigate
whether Raman spectroscopy can be used to a discriminate of egg quality.
Raman spectroscopy is a promising tool for detection of virus infection in live cells. In the present study, we
demonstrate its feasibility to observe dynamic reaction of the live cell infected by virus. The Raman spectra of the
adenovirus infected live cell (293 HEK) are analyzed by comparing with those of control cells. Principal
component analysis (PCA) is employed also to analyze the spectra in detail. A band at 1650 cm-1 increases its
intensity in the spectra measured at 24 hours after the virus infection. The infection of the virus is also examined
by immune-staining and transmission electron microscope (TEM), and the virus infection is confirmed with these
method also. It should be noted that the present technique does not require specifying the type of virus in
advance.
Live subcutaneous tumor grown in nude mouse is studied in situ with hyperspectral autofluorescence imaging and Raman spectroscopy. The purpose of the study is to develop methods for characterization of biochemical changing and of histological type of tumor without labeling. The results show that there are site depending variation in the fluorescence and Raman spectra. At the spot in which calcification is in process, Raman spectra showed a strong and specific band at 957 cm-1 due to PO4 species. The autofluosescence image can prove the histological changes based on the NADH and FAD which are major fluorophores in biological tissues. The hyperspectral image is analyzed with principal component analysis and the reconstructed images successfully depicts a different between necrotic and viable part within living subcutaneous tumor.
The esophageal cancer has a tendency to transfer to another part of the body and the surgical operation itself sometimes gives high risk in vital function because many delicate organs exist near the esophagus. So the esophageal cancer is a disease with a high mortality. So, in order to lead a higher survival rate five years after the cancer’s treatment, the investigation of the diagnosis methods or techniques of the cancer in an early stage and support the therapy are required. In this study, we performed the ex vivo experiments to obtain the Raman spectra from normal and early-stage tumor (stage-0) human esophageal sample by using Raman spectroscopy. The Raman spectra are collected by the homemade Raman spectrometer with the wavelength of 785 nm and Raman probe with 600-um-diameter. The principal component analysis (PCA) is performed after collection of spectra to recognize which materials changed in normal part and cancerous pert. After that, the linear discriminant analysis (LDA) is performed to predict the tissue type. The result of PCA indicates that the tumor tissue is associated with a decrease in tryptophan concentration. Furthermore, we can predict the tissue type with 80% accuracy by LDA which model is made by tryptophan bands.
The aim of the present study is to evaluate the capability of a miniaturized Raman endoscope (mRE) system to monitor the advancement of colorectal tumors in live model mice. The endoscope is narrow enough to observe the inside of the mouse colon under anesthesia. The mRE system allows to observe the tissues and to apply a miniaturized Raman probe for the measurement at any targeted point within the colon. Raman spectroscopy allows obtaining information about molecular composition without damaging the tissue (i.e., noninvasively). Continuous monitoring of the same tumor is carried out to study molecular alterations along with its advancement. The Raman spectra measured before and after the anticancer drug (5-FU) treatment indicated spectral changes in the tumor tissue. It suggests that the tumor is not cured but supposedly transformed to another tumor type after the treatment.
Our Raman probe that is called as ball-lens hollow fiber Raman probe (BHRP) had been proved
possessing capability to detect the biochemical alteration within biological tissue. Whether BHRP
has high capability and sensitivity in diagnosing the biochemical changing of tissue or not, mouse's
normal rectal and anorectal prolapse (AP) were decided to be used as a model for this non invasive
method. This AP is azoxymethane and DSS-induced mouse’s anorectal prolapse. Main outcome of
BHRP will be potential for non-invasive method in tumor diagnosing. BHRP spectra obtained were
a high quality and allowed analysis of their differences between normal rectal (control group) and
AP. After spectral acquisition and comparison with corresponding images of hematoxylin/eosinstained
section observation used to make the histopathologic diagnosing, BHRP detected some
differences within the region of moiety of DNA, protein (i.e. collagen) and lipid, then following with
the alteration of symmetric P=O stretching vibration compared with the normal rectal tissue. BHRP
discriminate normal tissue and AP in the real-time.
The esophageal cancer is a disease with a high mortality. In order to lead a higher survival rate five years after the
cancer’s treatment, we inevitably need a method to diagnose the cancer in an early stage and support the therapy. Raman spectroscopy is one of the most powerful techniques for the purpose. In the present study, we apply Raman spectroscopy to obtain ex vivo spectra of normal and early tumor human esophageal sample. The result of principal component analysis indicates that the tumor tissue is associated with a decrease in tryptophan concentration. Furthermore, we can predict the tissue type with 80% accuracy by linear discriminant analysis which model is made by tryptophan bands.
Problem of viruses is very actual for nowadays. Some viruses, which are responsible for human of all tumors, are about
15 %. Main purposes this study, early detection virus in live cell without labeling and in the real time by Raman
spectroscopy. Micro Raman spectroscopy (mRs) is a technique that uses a Raman spectrometer to measure the spectra of microscopic samples. According to the Raman spectroscopy, it becomes possible to study the metabolites of a live
cultured cell without labeling. We used mRs to detect the virus via HEK 293 cell line-infected adenovirus. We obtained
raman specters of lives cells with viruses in 24 hours and 7 days after the infection. As the result, there is some
biochemical changing after the treatment of cell with virus. One of biochemical alteration is at 1081 cm-1. For the clarification result, we use confocal fluorescent microscopy and transmission electron microscopy (TEM).
Ball-lens hollow fiber Raman Probe (BHRP) and FTIR spectroscopy were main tools in this study. Thus, both of
equipments detected the alteration of antisymmetric and symmetric P=O stretching vibration within our mice colorectal
tumor models. Some differences of spectra due to randomly the edge of each BHRP and FTIR attached the surface of
tumor during measurements. Meanwhile, the application of FTIR potentially differentiates the grade levels of non-clinic
samples colorectal tumor models at four different grades (normal, grade 1, grade 2 and grade 3). Detailed investigations
were assignable to wave numbers that publicized to represent biochemical alteration. The whole of investigated spectra
in the fingerprint region revealed some different peaks and shoulders, most of which were assignable to wave numbers
that exposed to represent biochemical alteration within the tissue. Differences in peak heights and peak ratio indicated
differences in biochemical composition of cancer from different grade level. However, all collected colorectal tumor
model at different peak was distinguishable, where antisymmetric and symmetric P=O stretching vibration was imaged
and mapped clearly by both equipments. Therefore, BHRP were comfortable for in vivo studies. Meanwhile FTIR
spectral analysis in combination with calibration curve might be used to distinguish cancer grade within colorectal tumor
model tissue for ex vivo study.
The CARS spectroscopy system using the dual-wavelength oscillation electronically wavelength tuned laser as a
pumping light source were constructed to realizes more sensitive full range non-chromosomal spectroscopy. Simpler
configuration of CARS optical system was realized by using the laser. To realize the CARS system, the methods to
synchronize the pulses with two different wavelengths generated from the dual-wavelength oscillation electronically
wavelength tuned laser was demonstrated by using sum frequency generation.
Raman spectroscopy is a powerful tool for analysis of the chemical composition in living tissue and cells without
destructive processes such as fixation, immunostaining, and fluorescence labeling. Raman microspectroscopic technique
enables us to obtain a high quality spectrum from a single living cell. We demonstrated in situ cell cycle analysis with
Raman microspectroscopy with the excitation wavelength of 532 nm. Cell cycle phases, G0/G1 and G2/M were able to
be identified in the present study. The result of in situ Raman analysis was evaluated with flow cytometry analysis.
Although the Raman spectra of living cells showed complex patterns during cell cycle, several Raman bands could be
useful as markers for the cell cycle identification. A single cell analysis using Raman microspectroscopy predicted a
possibility to observe directly molecular dynamics intracellular molecules of proteins, lipids and nucleic acids. Our
current study focused on cytoplasm region and resonant Raman signals of cytochrome c in mitochondrion, and discussed
how the Raman signals from cellular components contribute to the Raman spectral changes in cell cycle change in the
human living cell (lung cancer cell).
The Raman spectroscopic technique enables the observation of intracellular molecules without fixation or labeling procedures in situ. Raman spectroscopy is a promising technology for diagnosing cancers-especially lung cancer, one of the most common cancers in humans-and other diseases. The purpose of this study was to find an effective marker for the identification of cancer cells and their malignancy using Raman spectroscopy. We demonstrate a classification of cultured human lung cancer cells using Raman spectroscopy, principal component analysis (PCA), and linear discrimination analysis (LDA). Raman spectra of single, normal lung cells, along with four cancer cells with different pathological types, were successfully obtained with an excitation laser at 532 nm. The strong appearance of bands due to cytochrome c (cyt-c) indicates that spectra are resonant and enhanced via the Q-band near 550 nm with excitation light. The PCA loading plot suggests a large contribution of cyt-c in discriminating normal cells from cancer cells. The PCA results reflect the nature of the original cancer, such as its histological type and malignancy. The five cells were successfully discriminated by the LDA.
One of suspect environmental endocrine disruptors that affect mouse male reproduction by altering the morphology of Sertoli cells and spermatogenic cells is phthalate. The effects of mono(2-ethylhexyl)phthalate (MEHP), one of metabolites of di(2-ethylhexyl)phthalate , on immature mouse testes in vivo were examined. We have recently shown
that MEHP induced Sertoli cells necrosis and spermatogenic cells apoptosis in mice by TUNEL method, F-actin staining,
and ultrastructural study, but there is no data for biochemical changing of testes due to those methods could not explore.
To verify in detail of it, we conducted Raman spectroscopy study with 785 nm wavelength laser line, 50mW of laser power and 3 minutes of exposure time to analysis the MEHP-treated testicular tissue, which has been fixatived by 4% paraformaldehyde (PFA). Five weeks old (5 w.o) male mice were used in this experiment. As the results, the alterations were observed by Raman spectroscopy that there are significantly differences of DNA, actin filament, type IV collagen and amide I between control group (0 μM MEHP) and treatment group (100 μM MEHP). These results significantly support histology staining observation (such as the apoptotic spermatogenic cells which is associated with DNA fragmentation and F-actin disruption) and ultrastructural observation (such as mitochondria rupture and disintegration of nucleus membrane). Raman spectroscopy can be used for 4% PFA-fixatived tissue observation. However, we recommend that Raman spectroscopy may be able to be expanded as an armamentarium not just for the clarification of histology staining and ultrastructural study, but furthermore, it may be as a non-invasion assessment for screening animal tissue toxicity of chemical in future.
An optical biopsy system which equips miniaturized Raman probes, a miniaturized endoscope and a fluorescent image
probe has been developed for in vivo studies of live experimental animals. The present report describes basic optical
properties of the system and its application studies for in vivo cancer model animals and ex vivo human cancer tissues. It
was developed two types of miniaturized Raman probes, micro Raman probe (MRP) made of optical fibers and ball lens hollow optical fiber Raman probe (BHRP) made of single hollow optical fiber (HOF) with a ball lens. The former has rather large working distance (WD), up to one millimeter. The latter has small WD (~300μm) which depends on the focal length of the ball lens. Use of multiple probes with different WD allows one to obtain detailed information of subsurface tissues in the totally noninvasive manner. The probe is enough narrow to be inserted into a biopsy needle (~19G), for observations of the lesion at deeper inside bodies. The miniaturized endoscope has been applied to observe progression of a stomach cancer in the same rat lesion. It was succeeded to visualize structure of non-stained cancer tissue in live model animals by the fluorescent image technique. The system was also applied to ex vivo studies of human breast and stomach cancers.
The optical property of the ball lens mounted hollow optical fiber Raman probe (BHRP) is studied in the present study.
Since the ball lens has rather large aberration, the focus of the BHRP is dispersed and the spatial resolution in depth
direction goes low. The spatial dispersion of the focal point was evaluated using model samples. The BHRP equipped a
sapphire ball lens of 500 μm diameter was employed. Layered samples consisting of a polymethyl methacrylate
(PMMA) substrate and various thicknesses of polyethylene (PE) films were measured with the BHRP. The relative band
intensities of the upper and the lower layers appear at different rates in the obtained spectra, reflecting the optical
properties of the probe. According to the spectra, the optical dispersion of the focal point is estimated. The result
suggests that the spatial dispersion of the focus point fitted to Gaussian distribution. The working distance (WD) is 53
μm and the FWHM of the fitted Gauss distribution is 64 μm.
Living pancreatic cancer tissues grown subcutaneously in nude mice are studied by in vivo Raman spectroscopy and
autofluorescence imaging. Comparing the same point spectra of alive pancreatic cancer tissue to that of the dead tissue, it
is found that they are different each other. The results suggest that the spectral changes reflect the protein conformational
changes in the tumor tissue with death of the host animal. From the result of autofluorescence study, in vivo
autofluorescence imaging has potential as a method to assign the histological elements of the pancreatic cancer tissue
without any staining. These results strongly suggest that combination of these techniques is very important to study
biological tissue.
A Raman spectroscopic technique enables to observe intracellular molecules without fixation or labeling procedures in
situ. We demonstrated a classification of human lung cancer cells with Raman spectroscopy and principal component
analysis. Normal lung cell-lines and 4 pathological types of cancer cell-lines were seeded on culture dishes and
examined. It was as a preliminary study for direct Raman imaging spectroscopy, which could be available for clinical use,
to diagnose cancer. The result suggests that Raman spectroscopy could be a complementary method for
immunohistology study. We also constructed a new direct Raman imaging system consisting of a high sensitive CCD
image sensor, narrow band pass-filters, and a background-free electrically tunable Ti:Sapphire laser. The observation
wavelengths can be switched immediately for the purpose of malignancy rapid diagnosis or real time measurement for
apoptotic cells. The potential ability of the direct Raman imaging system is supposed to evaluate apoptosis by UV
irradiation and anticancer drug-treatment for living lung cancer cells.
Raman imaging methods provides visual information of pathological changes of tissues and cells without labeling procedures. We are developing direct Raman imaging technique in order to measure a Raman image of living tissues and cells in a short period without mapping procedure. A preliminary system is prepared consisting of a background-free tunable laser, band-pass filters and a CCD detector. In the present study, it is demonstrated the viability of the technique in a real time measurement of the direct Raman images.
Living pancreatic cancer tissues grown subcutaneously in nude mice are studied by in vivo microscope Raman
spectroscopy. Comparing the spectra of living pancreatic cancer tissue to that of the dead same tissue, it is found that
they are different each other. In the subtraction spectrum, Raman bands observed at 937, 1251, 1447 and 1671 cm-1 are
appeared in negative direction and those observed at 966 and 1045 cm-1 are appeared in positive direction. The results
strongly suggest that the spectral changes reflect the protein conformational changes in the tumor tissue with death of the
host animal. The present result demonstrates the importance of in vivo, real time studies of biomedical tissues using
Raman spectroscopy.
An optical biopsy system for small experimental animals has been developed. The system includes endoscope probe, portable probe and two kinds of miniaturized Raman probes. The micro Raman probe (MRP) is made of optical fibers and the ball lens hollow optical fiber Raman probe (BHRP) is made of hollow fiber. The former has large focal depth and suitable to measure average spectra of subsurface tissue. The latter has rather small focal depth and it is possible to control focal length by selecting ball lens attached at the probe head. It is suitable to survey materials at the fixed depth in the tissue. The system is applied to study various small animal cancer models, such as esophagus and stomach rat models and subcutaneous mouse models of pancreatic cancers. In the studies of subcutaneous tumor model mouse, it is suggested that protein conformational changes occur in the tumor tissue within few minutes after euthanasia of the mouse. No more change is observed for the following ten minutes. Any alterations in the molecular level are not observed in normal skin, muscle tissues. Since the change completes in such a short time, it is suggested that this phenomenon caused by termination of blood circulation.
Two types of hollow fiber-optic probes are developed to measure the in vivo Raman spectra of small animals. One is
the minimized probe which is end-sealed with the micro-ball lens. The measured spectra reflect the information of the
sample's sub-surface. This probe is used for the measurement of the esophagus and the stomach via an endoscope.
The other probe is a confocal Raman probe which consists of a single fiber and a lens system. It is integrated into the
handheld microscope. A simple and small multimodal probe is realized because the hollow optical fiber requires no
optical filters. The performance of each probe is examined and the effectiveness of these probes for in vivo Raman
spectroscopy is shown by animal tests.
A hollow optical fiber Raman probe equipped with a ball lens is developed to study the stomach cancer. The probe generates little Raman scattering or fluorescence noise because the HOF confine light into the air core. In order to maximize the collection efficiency and the spatial resolution, material and size of the ball lens is optimized. The total diameter of the probe head is 0.64 mm. The probe is useful for measurement of layered sample by the contact mode because it has very short focal length and high depth resolution. In vivo experiments in a stomach of a living rat demonstrate the high viability of the probe for noninvasive diagnosis of the stomach cancer.
Development of the optical biopsy system for experimental small animals is in progress. A prototype of the system which consists of a miniaturized gastro endoscope unit and Raman probes has been completed by now. The system is developed to study a gastric cancer rat model. The endoscope is 2.5 mm in diameter and is equipped an imaging bundle fiber, illumination fibers, a channel and a mechanism to angle the probe head. The head of the Raman probe comes out through the channel and it is possible to aim the probe to the target watching on the monitor. The endoscope was inserted into the anaesthetized healthy rat under the breathing support. It was successfully observed inside of the stomach of the living rat and measured Raman spectra. The spectrum of blood vessels contains the strong contribution from lipids. The present results demonstrate high potential of the system in the in vivo Raman study using the rat model.
A confocal Raman probe based on a silver hollow optical fiber is developed to apply in remote optical biopsy via a micro-endoscope. The probe consists of a single hollow optical fiber delivering both the excitation and the Raman scattered light, and a confocal end unit attached at the distal end of the fiber. We demonstrate a high spatial resolution remote Raman measurement with the present probe. The measured spectrum shows a depth-resolution of below 0.15 μm. In addition, we propose a ball lens coupled miniature high-resolution Raman probe and show that it operates as well as the confocal probe.
We have developed an excitation wavelength tunable Raman system which consists of a spectrometer, a CCD detector and an electronically tuned Ti:sapphire (ETT) laser. All those components are controlled with a single computer. This system is useful for Raman excitation profile measurements. A new technique was developed to calibrate deviation of wavelengths of ETT laser from that of Raman spectrometer within +/- 1.5cm-1 of error. Obtained Raman spectra of hemoglobin measured with various excitation wavelengths in 700-860 nm range showed characteristic change of their intensities in some of Raman bands. Assignments of Raman bands were carried out sufficiently in help with change in intensity of the bands which reflect symmetric character of the responsible vibrational modes with the change in excitation wavelength.
A variety of progress concerned with an electronically tuned Ti:sapphire laser with an acousto-optic tunable filter for the purpose of spectroscopic applications are described. The major advances are that fast and random access tuning can be achieved in a tuning range of 690-1056 nm by computer control without any mechanical changes of cavity. The access speed reached to 250 microsecond(s) . The Ti:sapphire laser also provided tunable dual-wavelength operation in a single laser cavity with introducing two different radio frequencies at same time. The operation was promising as a pumping source of difference-frequency generation. Difference-frequency generation of non-mechanical tuning was realized from 6.0 to 7.1 micrometers , from 6.8 to 8.6 micrometers , and from 8.5 to 11.3 micrometers for the phase-matching angle at 58, 53, and 46 deg, respectively, using the dual-wavelength laser. Furthermore, the electronic tuning by acousto-optic tunable filter achieved broad tunable picosecond pulse generation with only computer control.
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