Continuous wave THz spectroscopy has been used to obtain spectra for four isostructural dipeptide nanotubes at 4.2K from 2 cm^-1 to 100 cm^-1 (0.05 to 3 THz). Line-narrowing of spectral features by a factor of 2 to 4 is observed for the crystalline dipeptide films investigated by absorption spectroscopy using a plane parallel waveguide, compared to spectra from pressed disks of polyethylene-diluted samples. The x-ray determined crystal structures of these peptides formed the basis for a parallel computational investigation. Spectral predictions from the ab initio level computational package DMOL³ and the empirical force field model CHARMM22 are compared to the experimentally obtained THz absorption spectra. The THz waveguide spectroscopy technique can provide information on the orientation-dependent dipole coupling of the vibrational modes, which can aid in validating computational models.

A Raman spectroscopic investigation of a genetically altered Human Embryonic Kidney Cell (HEK293) along with a pathologically normal cell has been carried out by a conventional method. The genetic alteration was carried out with a standard protocol by using a Green Fluorescence Protein (GFP). Raman spectra show that there are dramatic differences between the spectrum obtained from a genetically altered cell and that obtained from a pathologically normal cell. The former shows three broad bands; meanwhile the latter shows several sharp peaks corresponding to the ring vibrational modes of Phen, GFP and DNA. The present analysis provides an indication that the force field near Phen located at 64, 65 and 66 was altered during the genetic transformation. The Raman spectrum could be a direct experimental evidence for substantial modifications triggered due to the expression of specific genes.

Raman spectroscopy has proved to be a highly sensitive tool for differentiating between normal, cancerous or pre-cancerous tissues. To date, histological application of Raman mapping has been limited due to lengthy mapping times. Streamline^TM Raman imaging is a novel mapping technique that has reduced total mapping times to a level that is becoming clinically practicable. Raman Streamline mapping was carried out on a 20μm frozen section of an oesophageal biopsy. A contiguous 7μm section was stained with haematoxylin and eosin (H&E) with histpathology analysed by a pathologist. The step size and acquisition times were varied and the resulting spectra, principal component score maps and loads were compared. The signal to noise for the raw spectra and a relative 'signal to noise' of the principal component loads were determined. The Streamline mapping technique was also compared to traditional point Raman mapping. The principal component loads were similar despite varying the acquisition time and number of spectra, with the fifth load used for comparison of the noise levels. Gross biochemical information was extracted showing good correlation with the H&E section even for short overall mapping times as low as 30-90 minutes for a biopsy ~2mm in diameter (0.5s acquisition time per 25.3μm Raman pixel). Streamline mapping was of the order of 3-7 times faster than traditional point mapping with the greatest improvement made for high resolution maps. Further optimization of the system is still possible which will reduce this mapping time further making implementation in a clinical environment a future possibility.

Fourier transform infrared (FT-IR) spectroscopic imaging is an emerging technique that combines the molecular selectivity of spectroscopy with the spatial specificity of optical microscopy. We demonstrate a new concept in obtaining high fidelity data using commercial array detectors coupled to a microscope and Michelson interferometer. Next, we apply the developed technique to rapidly provide automated histopathologic information for breast cancer. Traditionally, disease diagnoses are based on optical examinations of stained tissue and involve a skilled recognition of morphological patterns of specific cell types (histopathology). Consequently, histopathologic determinations are a time consuming, subjective process with innate intra- and inter-operator variability. Utilizing endogenous molecular contrast inherent in vibrational spectra, specially designed tissue microarrays and pattern recognition of specific biochemical features, we report an integrated algorithm for automated classifications. The developed protocol is objective, statistically significant and, being compatible with current tissue processing procedures, holds potential for routine clinical diagnoses. We first demonstrate that the classification of tissue type (histology) can be accomplished in a manner that is robust and rigorous. Since data quality and classifier performance are linked, we quantify the relationship through our analysis model. Last, we demonstrate the application of the minimum noise fraction (MNF) transform to improve tissue segmentation.

Transient fluorescence detected infrared (TFD-IR) microscopy was developed to overcome the diffraction limit of infrared (IR) light without a near-field system. This microscopic technique is based on TFD-IR spectroscopy, which converts information on IR absorption to fluorescence intensity by further electronic excitation of vibrationally excited molecules by a probing UV/visible light. Roots of Arabidopsis thaliana and living A549 cells with fluorescent dyes were chosen as samples. In the measurements using the TFD-IR microscope, tunable IR picosecond laser pulses were used in the wavelength range from 2700 to 3700 nm, corresponding to CH, NH, and OH stretching modes. Fluorescence images of the root cells of A. thaliana by the TFD-IR scheme were obtained with super-resolution compared with the resolution of conventional IR microscopy. The resolution is estimated to be less than 2.6 μm by fitting of a gaussian function. However, the TFD-IR images were dominated mainly by the fluorescent dyes because they were almost the same as a conventional fluorescence image. To investigate other contributions hidden by that of fluorescent dyes, we plotted the fluorescence intensity in several 5 μm squares at various IR wavelengths, called a TFD-IR spectrum. For root cells of A. thaliana, the TFD-IR spectra show shapes similar to those of a conventional IR absorption spectrum of the fluorescent dye. Therefore, the TFD-IR images are not due to the cellular components. For an A549 cell, the TFD-IR spectra were different from a conventional IR absorption spectrum of fluorescent dyes in the wavelength region shorter than 3100 nm. We speculate that the spectral difference is due to the cellular components, possibly assigned to the combination band related to amino groups of cellular components bonded covalently to the fluorescent dyes.

In recent years, various types of molecular imaging technologies have been developed, but many of them require probes and may have some influence on the distribution of the target molecules. In contrast, Raman microscopic analysis is effective for molecular identification of materials, and molecular imaging methods employing Raman scattering light can be applied to living organisms without use of any exogenous probes. Unfortunately, Raman microscopic imaging is rarely used in the biomedical field due to the weakness of Raman signals. When the conventional Raman microscopes are used, the acquisition of an image of a cell usually takes several hours. Recently, a slit-scanning confocal Raman microscope has been developed. It can acquire images of living cells and tissues with faster scanning speed. In this study, we used the slit-scanning confocal Raman microscope (RAMAN-11) to image the distribution of a drug in living cells. We could acquire images of the distribution of an anticancer reagent in living cells within several minutes. Since the wavelength of Raman scattering light is determined by the frequency of molecular vibration, the in situ mapping of the intracellular drugs without use of a probe is possible, suggesting that laser Raman imaging is a useful method for a variety of pharmacokinetic studies.

This study attempts to combine the tear ferning test and the drop coating deposition Raman spectroscopy (DCDRS) technique to analyze the biochemical composition of human tear fluid from healthy volunteers. DCDRS has been shown to be a highly reproducible and sensitive method of obtaining Raman spectra from low concentration protein solutions making it ideal for the analysis of tear fluid. On drying, tear samples were found to produce ring-shaped patterns, which are characteristic of the DCDRS technique, with additional fern-like structures produced inside the rings. The biochemical composition of the each drying pattern was studied by Raman point mapping and principal components analysis. Assignment of high-signal-to-noise tear spectra showed that tear proteins, urea, bicarbonate and lipid components were all present in the dried tear drop. Comparing an image time series of the drying process with the biochemical distributions from the Raman point map revealed the order of biochemical deposition in the drying pattern. The combination of DCDRS and the tear ferning test shows enough promise to be further studied as a near-patient technique for assisting the diagnosis of ocular infection, but further work is required to validate the technique.

We have developed a compact portable instrument for resonance Raman spectroscopy of carotenoid molecules in skin tissue. Our application focuses on the 1525 cm^-1 Raman line common to all carotenoids. We use a divided shifted Raman spectroscopy (DSRS) technique that reduces sensitivity to detector drift and error. Two wavelength-narrowed LEDs illuminate the sample, and scattered light in four different wavelength channels is measured. This multi-spectral approach has single-photon sensitivity and compares favorably with laser-based Raman measurements in terms of accuracy, repeatability, and measurement time.

Monitoring the sterility of cell or tissue cultures is an essential task, particularly in the fields of regenerative medicine and tissue engineering when implanting cells into the human body. We present a system based on a commercially available microscope equipped with a microfluidic cell that prepares the particles found in the solution for analysis, a Raman-spectrometer attachment optimized for non-destructive, rapid recording of Raman spectra, and a data acquisition and analysis tool for identification of the particles. In contrast to conventional sterility testing in which samples are incubated over weeks, our system is able to analyze milliliters of supernatant or cell suspension within hours by filtering relevant particles and placing them on a Raman-friendly substrate in the microfluidic cell. Identification of critical particles via microscopic imaging and subsequent image analysis is carried out before micro-Raman analysis of those particles is then carried out with an excitation wavelength of 785 nm. The potential of this setup is demonstrated by results of artificial contamination of samples with a pool of bacteria, fungi, and spores: single-channel spectra of the critical particles are automatically baseline-corrected without using background data and classified via hierarchical cluster analysis, showing great promise for accurate and rapid detection and identification of contaminants.

The characterization and identification of microorganisms by infrared or Raman spectroscopy is probably one of the best developed and most frequent applications of biomedical vibrational spectroscopy. The serial types of dedicated instruments for routine FT-IR characterizations of microorganisms are now available on the market and already used in routine microbiological laboratories. The experiences gained to date, especially the necessity to define standards for sampling and measurement procedures and the details of how data compatibility between different laboratories is achieve will be discussed as well as the problem to establish validated reference data bases for objective species or strain identifications.

It has been less than three years since the beginnings of systematic development of fiber optic Raman illumination/collection systems for operation in the presence of extensive photon diffusion. These fiber optic probes depend on spatial separation of illumination and collection regions to deep Raman spectroscopy and imaging in highly scattering media. Both backscattering and transmission systems have been described in the literature. They are closely related to probes used for fluorescence imaging and tomography in turbid media, but in most there are modifications to allow distributed laser illumination to provide high absolute power without thermal damage. We review the history of the spatially resolved techniques, discuss data reduction issues and current probe designs and survey current applications to human and animal tissue.

Our previous results from Raman spectroscopy studies on ex vivo lung tissue showed the technique had great potential to differentiate between samples with different pathologies. In this work, a fast dispersive-type near-infrared (NIR) Raman spectroscopy system was developed to collect real-time, noninvasive, in vivo human lung spectra. The 785 nm excitation, and the collection of tissue emission were accomplished by using a reusable fiber optic catheter which passed down the instrument channel of a bronchoscope. Filters in two stages blocked laser emission other than 785 nm from reaching the tissue surface, and reduced fiber fluorescence and elastically scattered excitation light from being passed to the spectrometer. The spectrometer itself consisted of one of two holographic gratings with usable frequency ranges of: 700 to 2000 cm^-1 and 1500 to 3400 cm^-1. The dispersed light was detected by a cooled CCD array consisting of 400 by 1340 pixels. To increase the resolution of the system, while maximizing the throughput, a second fiber bundle, consisting of 54×100 μm diameter fibers connected the catheter to the spectrometer. The fibers in this second bundle were spread out to form a parabolic arc which replaced the conventional entrance slit. This geometry corrected for image aberrations, permitting complete CCD vertical binning, thereby yielding up to a 20-fold improvement in signal-to-noise ratio. The estimated spectral resolution of the system was 9 cm^-1 for both gratings. So far we have measured spectra from 20 patients and have seen clear differences between spectra from tumor and normal tissue.

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.

A program of work has been established to explore the possibility of sampling Raman information from material deeply buried within tissues. Time-gating, spatial offsetting of source and collection (spatially offset Raman spectroscopy) and transmission approaches have been explored. This invited contribution will outline these and the limitations of each technique will be discussed. Time-gating allows well defined depth selection, but with low penetrations of the order of 1- 2mm; whereas SORS has allowed some depth selection and an order of magnitude greater depth range; the use of transmission Raman spectroscopy has permitted greater depths of penetration, but depth selection is not possible in this configuration. To date transmission Raman has demonstrated recovery of compositional information through 27mm of mammalian tissue. Furthermore, a first demonstration of Kerr-gated Raman spectroscopy for fluorescence suppression in resonance Raman measurements of liver and kidney is also outlined.

Raman spectroscopy has been well established as a powerful method for studying biological tissues and diagnosing diseases. In this study we have developed a breast cancer animal model and collected in vivo Raman spectra of mammary glands of 27 Sprague-Dawley female rats treated with DMBA and 5 non-treated used as control group. A dispersive Raman spectrometer with a @785 nm laser excitation coupled a fiber optic probe and a CCD detector was used to obtain the spectra. The obtained in vivo transcutaneous Raman spectra have shown important differences between normal and abnormal tissues when acquired from one side to the other side of the lesion.

Early detection of malignant tumours, or their precursor lesions, can dramatically improve patient outcome. High risk human Papillomavirus (HPV), particularly HPV16, infection can lead to the initiation and development of uterine cervical neoplasia. Bearing this in mind the identification of the effects of HPV infection may have clinical value. In this manuscript we investigate the application of Raman microspectroscopy to detect the presence of HPV in cultured cells when compared with normal cells. We also investigate the effect of sample fixation, which is a common clinical practice, on the ability of Raman spectroscopy to detect the presence of HPV. Raman spectra were acquired from Primary Human Keratinocytes (PHK), PHK expressing the E7 gene of HPV 16 (PHK E7) and CaSki cells, an HPV16 containing cervical carcinoma derived cell line. The average Raman spectra display variations, mostly in peaks relating to DNA and proteins, consistent with HPV gene expression and the onset of neoplasia in both live and fixed samples. Principle component analysis was used to objectively discriminate between the cells types giving sensitivities up to 100% for the comparison between PHK and CaSki. These results show that Raman spectroscopy can discriminate between cell lines representing different stages of cervical neoplasia. Furthermore Raman spectroscopy was able to identify cells expressing the HPV 16 E7 gene suggesting the approach may be of value in clinical practice. Finally this technique was also able to detect the effects of the virus in fixed samples demonstrating the compatibility of this technique with current cervical screening methods. However if Raman spectroscopy is to make a significant impact in clinical practice the long acquisition times must be addressed. In this report we examine the potential for beam shaping and advanced to improve the signal to noise ration hence subsequently facilitating a reduction in acquisition time.

An overview is presented of recent trends in coherent anti-Stokes Raman scattering (CARS) microscopy. We briefly discuss the influence of tissue scattering on the CARS signal, methods for controlling the CARS emission and prospects for surface-enhancement of the CARS radiation.

A microscope system has been constructed that allows simultaneous acquisition of Raman scattering spectra and elastic scattering Fourier-plane data. The Raman scattering channel reports on chemical composition of the microscopic sample while the elastic scattering channel reports on morphological information about the sample. The system has been validated by acquiring data from single polystyrene beads and analyzing the elastic scattering signal using Generalized Lorenz-Mie Theory while comparing the Raman scattering signature to other polystyrene spectra from the literature. Monocytes and neutrophils, two immune cell types, have also been studied and show clear chemical and morphological differences between cell types.

We report tomographic reconstruction of objects located several millimeters below the surface of highly scattering media. For this purpose we adapted proven software developed for fluorescence tomography with and without the use of spatial priors¹. For this first demonstration we acquired Raman spectra using an existing ring/disk fiber optic probe with fifty collection fibers². Several illumination ring diameters were employed to generate multiple angles of incidence. Tomographic reconstruction from Raman scatter was tested using a 9.5 mm diameter Teflon® sphere embedded in a gel of agarose and 1% Intralipid. Blind reconstruction of the sphere using the 732 cm^-1 C-F stretch yielded an accurate shape but an inaccurate depth. Using the known shape and position of the object as spatial priors, a more accurate reconstruction was obtained. We also demonstrated a reconstruction of the tibial diaphysis of an intact canine hind limb using spatial priors generated from micro-computed tomography. In this first demonstration of Raman tomography in animal tissue, the P-O stretch of the bone mineral at 958 cm^-1 was used for the reconstruction. An accurate shape and depth were recovered.

Raman spectroscopic studies have shown that the properties of the organic matrix and the orientation of the mineral and matrix components of bone have a large influence on its properties. We employ polarized Raman microspectroscopy to monitor the changes in the orientation of mineral crystallites during tensile loading of bovine femora in the elastic regime. We load tissue in a custom-built dynamic mechanical tester that fits on the stage of a Raman microprobe and can accept hydrated tissue specimens. Parallel and perpendicular polarization components of the Raman spectra along the long axis of the diaphysis are obtained. We propose that the orientation and structure of mineral crystallites change on deformation of bone tissue by tensile loading.

We report the experimental results of physiologically active substances including hormones utilizing Terahertz (THz) spectrum technology. Various analytical techniques have been employed for the determination of physiologically active substances. Most of methods used oxidation reagent and sodium sulfite stabilization reagent or fluorescent reagent. These methods had a good selectivity, but the stability of samples was not satisfactory. The direct detection method has not been advanced yet. In this study, the THz characteristics of physiologically active substances were measured directly. We found all of samples have their vibrational features like signature peaks either in pellet and/or sample solution. A membrane device was used to hold the sample solution in this study. This device allows samples to be prepared in solutions and measured easily with THz measurement system after dried. Results suggest that this membrane device is sensitive for detecting the physiologically active substances in THz ranges. THz spectrum technology has the potential to be a useful tool in clinical applications. This approach promotes the understanding of the relationships between biomolecules with THz radiation.

Human synovial fluid droplets were investigated using drop deposition in combination with Raman spectroscopy. Following informed consent, synovial fluid was obtained from forty human patients with various severities of knee pain and/or osteoarthritis at the time of knee arthroscopy or total joint replacement. Synovial fluid was aspirated from the knee joint of each patient and stored at -80°C until examination by near-infrared Raman spectroscopy. Synovial fluid aspirates from the knee joint of each patient were deposited onto a clean fused silica microscope slide and the droplet dried under ambient laboratory conditions. Each droplet was illuminated by a line-focused or a ring-focused 785 nm laser. As the droplet dries, biofluid components segregated based on solubility differences and a deposit that is spatially heterogeneous was made. Spectra taken from the droplet edges and center were dominated by protein bands and showed the presence of at least two protein moieties in the droplet. Band area and band height ratios (1410 cm^-1/1450 cm^-1) showed the greatest change between specimens from patients with mild/early osteoarthritis compared to those with severe/late stage osteoarthritis. The greatest differences were found in the center of the droplet, which contains more soluble protein components than the edges.

Understanding compositional changes that occur when bone fails may help predict fracture risk. Compositional differences that arise among failed, strained, and undamaged regions of bone can be determined using Raman spectroscopy and double-notch specimens. A double-notch specimen is a rectangular bone beam that has identical, rounded notches milled equidistant from each end. When subjected to a four-point bend test, maximum strains occur at the roots of the notches, and eventually the bone fractures at one of the notches. Because both notches experience the same force, when one notch breaks, the other is 'frozen' in the state directly preceding fracture. Spectra taken at the roots of both the unbroken and fractured notches can measure changes in tissue that occur prior to and after bone failure, respectively. Phosphate center of gravities (CGs) were calculated and compared among three regions: control, strained (root of unbroken notch), and failed (root of fractured notch). In comparison to control regions, the phosphate CGs near the unbroken notch showed a shift toward higher wavenumbers ( > 0.5 cm^-1), with the shift being concentrated at the corners of the notch. The tissue in the failed region appears to have relaxed, and showed a shift toward higher wavenumbers ( > 0.5 cm^-1) only near the edge of the fracture.

The histogenesis of the breast Paget's disease was investigated by the optical diagnosis technique using Raman spectroscopy. A total of 15 spectra of the associated breast lesion, 21 spectra of the eczematoid skin lesion and 396 spectra of invasive breast cancer not otherwise specified were compared by clustering the spectral data between 800 - 1800 cm^-1 at level of similarity of 95%, using a correlation distance measurement by computing the covariance matrix. The Raman spectral-biochemical characterization of invasive breast cancer and breast Paget's disease with eczematoid skin lesion associated with underlying invasive breast lesion tissues enabled one concludes that the parenchymal disease had similar characteristics to the skin's Paget lesion. This could indicate a similar histogenesis for both. Thus, the findings of the present work adds a relevant experimental evidence that agrees with the epidermotropic theory of Paget's disease, that states that the cells originate in the breast ducts and migrate to the nipple's skin.

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.

The oxidative stress due to free radicals is implicated in the pathogenesis of tissue damage in diseases such as muscular dystrophy, Alzheimer dementia, diabetes mellitus, and mitochrondrial myopathies. In this study, the acute oxidative stress induced changes in nicotinamide adenine dinucleotides in mouse skeletal muscles are studied in vitro using Raman spectroscopy. Mammalian skeletal muscles are rich in nicotinamide adenine dinucleotides in both reduced (NADH) and oxidized (NAD) states, as they are sites of aerobic and anaerobic respiration. The relative levels of NAD and NADH are altered in certain physiological and pathological conditions of skeletal muscles. In this study, near infrared Raman spectroscopy is used to identify the molecular fingerprints of NAD and NADH in five-week-old mice biceps femoris muscles. A Raman vibrational mode of NADH is identified in fresh skeletal muscle samples suspended in buffered normal saline. In the same samples, when treated with 1% H₂O₂ for 5 minutes and 15 minutes, the Raman spectrum shows molecular fingerprints specific to NAD and the disappearance of NADH vibrational bands. The NAD bands after 15 minutes were more intense than after 5 minutes. Since NADH fluoresces and NAD does not, fluorescence spectroscopy is used to confirm the results of the Raman measurements. Fluorescence spectra exhibit an emission peak at 460 nm, corresponding to NADH emission wavelength in fresh muscle samples; while the H₂O₂ treated muscle samples do not exhibit NADH fluorescence. Raman spectroscopy may be used to develop a minimally invasive, in vivo optical biopsy method to measure the relative NAD and NADH levels in muscle tissues. This may help to detect diseases of muscle, including mitochondrial myopathies and muscular dystrophies.

The purpose of this study is to explore the feasibility of utilizing near-infrared (NIR) autofluorescence spectroscopy for in vivo diagnosis of precancer (i.e., dysplasia) in the cervix. A rapid NIR spectroscopy system in combination with a fiber-optic probe was developed for the in vivo NIR fluorescence measurements under the 785 nm laser excitation. Multivariate statistical techniques including principal component analysis (PCA) and linear discriminant analysis (LDA) were employed to develop the diagnostic algorithms for spectra classification. Classification result obtained from the PCA-LDA model based on tissue NIR autofluorescence data yielded a diagnostic sensitivity of 84.8% and specificity of 85.1% for discrimination of precancer from normal cervical sites. The results demonstrate that NIR autofluorescence technique has the capacity for the noninvasive, in vivo diagnosis of precancer in the cervix.

Optical properties of near-surface kidney tissue were monitored in order to assess response during reperfusion to long (20 minutes) versus prolonged (150 minutes) ischemia in an in vivo rat model. Specifically, autofluorescence images of the exposed surfaces of both the normal and the ischemic kidneys were acquired during both injury and reperfusion alternately under 355 nm and 266 nm excitations. The temporal profile of the emission of the injured kidney during the reperfusion phase under 355 nm excitation was normalized to that under 266 nm as a means to account for changes in tissue optical properties independent of ischemia as well as changes in the illumination/collection geometrical parameters in future clinical implementation of this technique using a hand-held probe. The scattered excitation light signal was also evaluated as a reference signal and found to be inadequate. Characteristic time constants were extracted using a fit to a relaxation model and found to have larger mean values following 150 minutes of injury. The mean values were then compared with the outcome of a chronic survival study where the control kidney had been removed. Rat kidneys exhibiting longer time constants were much more likely to fail. This may lead to a method to assess kidney viability and predict its ability to recover in the initial period following transplantation or resuscitation.

This study aims towards applying the intrinsic polarized fluorescence technique for obtaining valuable information from human cervical tissue samples. The efficacy of this technique is tested in human tissues by comparing its diagnostic capabilities with the bulk fluorescence. It is seen that biochemical information is hidden due to the presence of distortions by tissue scattering and absorption in the fluorescence spectra. Intrinsic tissue fluorescence provides a complete understanding of the biochemical and/or morphological changes that take place during the progression of disease. Recording of the experimental data and thereafter subsequent extraction of the intrinsic fluorescence serves as fingerprints towards determining the occurrence of these diseases. Here we report a comparative study of intrinsic versus bulk polarized fluorescence in cervical tissues. Intrinsic fluorescence is seen to be a more sensitive technique than bulk fluorescence for diagnosis of cervical cancers. Attempts have been made to study the changes in the amount of different fluorophores found in the epithelial and stromal layer of cervical tissue (both normal and cancerous). It has been seen that collagen decreases and NADH increases as a healthy cervical tissue develops into a cancerous one. Intrinsic fluorescence provides more consistent discriminating results as compared to bulk polarized fluorescence. It is also more sensitive in giving biochemical information from the different layers in cervical tissue. It may be concluded that intrinsic fluorescence shows promise as a viable tool for providing valuable insights towards fruitful diagnosis of the various stages of disease development and changes occurring with age.

In normal cell the mitochondria are the major source of energy for cellular functions. They serve as biosensors for oxidative stress and involved also in termination of cell function by apoptosis. The involvement of mitochondria in pathological states such as neurodegenerative diseases, sepsis, stroke and cancer are well documented. The involvement of mitochondrial respiration and function in cancer development, proliferation and possible therapy were initiated 75 years ago by Otto Warburg. Monitoring of NADH fluorescence in vivo as an intracellular oxygen indicator was established in the 1950-1970 by Britton Chance and collaborators. In the last 20 years we developed and used a multiparametric monitoring system enabling real time assessment of mitochondria NADH, microcirculatory blood flow and volume as well as HbO₂ oxygenation. In order to use this technology in clinical practice the commercial developed device-the "CritiView" was tested in animal models as well as in patients hospitalized in the critical care departments. In patients we tested the viability of the urethral wall (a less-vital tissue) by a 3 way Foley urinary catheter that contains the optical probe. The catheter was introduced to patients underwent open heart by-pass surgery or abdominal aorta aneurysm (AAA) operations. The monitoring started immediately after the insertion of the catheter to the patient and was stopped when the patient was discharged from the operation room. The results show that monitoring of the vitality of the Urethral wall provides information in correlation to the surgical procedure performed. In the AAA patients the occlusion of the aorta led to severe ischemia developed in the urethral wall and recovery of signals were recorded after the reopening of the aorta. In patients under went heart bypass surgery the urethra vitality was decreased dramatically during the operation and recovery was noted in most patients after the discharge of the patient from the operation room.

We report an ex-vivo study on Raman spectra of adipose tissue covered by layers of aortic intimal wall tissue with different thicknesses. The Raman vibration modes of 1435cm^-1, 2850cm^-1 and 2892cm^-1 were investigated for the first time on fresh porcine aortic adipose tissue with 633 nm laser excitation. The adipose tissue was taken from adventitial fat grown on aorta walls. The frozen sections of porcine aorta wall tissue with a thickness of 25μ - 50μ were cut from intimal surface. The samples were prepared by placing the variable numbers of the aorta intimal wall tissue layers on the top of adipose tissue. The Raman spectra of adipose tissue were investigated. The changes of intensities of the Raman modes versus thickness of the aorta intimal wall tissue layers were measured. The total thickness of the aorta intimal wall tissue layers was varied in the range of 50μ - 1800μ. The main characteristic Raman vibration modes of adipose tissue were found at 1435cm^-1, 2850cm^-1 and 2892cm^-1. Among them, the intensities of the modes of 2850 cm^-1 and 2892 cm^-1 are about four-times stronger than that of the 1435cm^-1 mode. The study on Raman vibration modes of 1435cm^-1, 2800 cm^-1 and 2950 cm^-1 may be useful for developing a simple, inexpensive and accurate optical technique for monitoring the degree of vulnerability of the aorta intimal surface due to atherogenesis. These three Raman modes can be used as new molecular spectroscopic indicators to monitor in situ the development of fatty-streaks and lipid core in aorta walls, and determine the thickness change of aorta intimal wall at different stages of atherogenesis.

The statistical and characteristic features of the polarized fluorescence spectra from cancer, normal and benign human breast tissues are studied through wavelet transform and singular value decomposition. The discrete wavelets enabled one to isolate high and low frequency spectral fluctuations, which revealed substantial randomization in the cancerous tissues, not present in the normal cases. In particular, the fluctuations fitted well with a Gaussian distribution for the cancerous tissues in the perpendicular component. One finds non-Gaussian behavior for normal and benign tissues' spectral variations. The study of the difference of intensities in parallel and perpendicular channels, which is free from the diffusive component, revealed weak fluorescence activity in the 630nm domain, for the cancerous tissues. This may be ascribable to porphyrin emission. The role of both scatterers and fluorophores in the observed minor intensity peak for the cancer case is experimentally confirmed through tissue-phantom experiments. Continuous Morlet wavelet also highlighted this domain for the cancerous tissue fluorescence spectra. Correlation in the spectral fluctuation is further studied in different tissue types through singular value decomposition. Apart from identifying different domains of spectral activity for diseased and non-diseased tissues, we found random matrix support for the spectral fluctuations. The small eigenvalues of the perpendicular polarized fluorescence spectra of cancerous tissues fitted remarkably well with random matrix prediction for Gaussian random variables, confirming our observations about spectral fluctuations in the wavelet domain.

Autofluorescence endoscopy is a promising modality for diagnosis of colonic tumors. This article discusses the origin of autofluorescence of the normal colon. Excised normal colons were analyzed by using fluorescence stereomicroscopy and a fluorescence-lifetime microscopy system. Fluorescence images showed that the mucosa had stronger autofluorescence than the submucosa. The results of fluorescence-lifetime measurement showed that nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) might be responsible for the autofluorescence of the colonic epithelia. Our results suggest that the mucosal autofluorescence generates by NADH and FAD was an important source of the green autofluorescence.