Diffuse reflectance spectroscopy of human adenomatous colon polyps in vivo

Diffuse reflectance spectra were collected from adenomatous colon polyps (cancer precursors) and normal colonic mucosa of patients undergoing colonoscopy. We analyzed the data by using an analytical light diffusion model, which was tested and validated on a physical tissue model composed of polystyrene beads and hemoglobin. Four parameters were obtained: hemoglobin concentration, hemoglobin oxygen saturation, effective scatterer density, and effective scatterer size. Normal and adenomatous tissue sites exhibited differences in hemoglobin concentration and, on average, in effective scatterer size, which were in general agreement with other studies that employ standard methods. These results suggest that diffuse reflectance can be used to obtain tissue information about tissue structure and composition in vivo.     

In vivo measurements of the wavelength dependence of tissue-scattering coefficients between 760 and 900 nm measured with time-resolved spectroscopy

We present in vivo values for the optical transport coefficients (mu(a), mu(s)?) of the adult human forearm, calf, and head from 760 to 900 nm measured with time-resolved spectroscopy. The accuracy of the method is tested with tissue-simulating phantoms. We obtain mu(s)?(lambda) approximately 1.1 - (5.1 x 10(-4) lambda) mm(-1) (forearm), 1.6 - (8.9 x 10(-4) lambda) mm(-1) (calf), and 1.45 - (6.5 x 10(-4) lambda) mm(-1) (head), where lambda is measured in nanometers. At 800 nm we obtain mu(a) = 0.023 +/- 0.004 mm(-1) (forearm), 0.017 +/- 0.005 mm(-1) (calf), and 0.016 +/- 0.001 mm(-1) (head). Our values differ substantially from published in vitro data. In particular, our transport coefficients for the adult head are substantially lower than previously reported values for adult human cerebral matter and pig skull cortical bone measured in vitro.     

In situ time-resolved fluorescence spectroscopy in the frequency domain in capillary electrochromatography

In situ time-resolved fluorescence spectroscopy for capillary electrochromatography (CEC) is described in the frequency domain. Fluorescence decay of the solute molecules is collected directly in the packed stationary phase of the CEC capillary. The fluorescence lifetime profile of the solute molecules reveals the microenvironments they experience in the C18 chromatographic interface. A quartz flow cell and experimental optimization of the signal-to-noise ratio are described that enable the collection of high-quality decay data and subsequent calculation of fluorescence lifetime profiles of the solute molecules. The distribution of pyrene (PY), 1-pyrenemethanol (PY-MeOH), and 1-pyrenebutanol (PY-BuOH) into the C18 stationary phase and the solute-C18 phase interactions are probed, under separation conditions for CEC. All three molecules display a Gaussian distribution of lifetimes, consistent with an ensemble of heterogeneous microenvironments in the C18 stationary phase. The least polar molecule PY diffuses deeply into and interacts extensively with the C18 phase, experiencing high hydrophobicity and significant heterogeneity of microenvironments. The retention order of PY-MeOH, PY-BuOH, and PY in CEC is determined by their interactions with the stationary phase, revealed by their fluorescence lifetime distributions.     

Investigation of two-layered turbid media with time-resolved reflectance

Light propagation in two-layered turbid media that have an infinitely thick second layer is investigated with time-resolved reflectance. We used a solution of the diffusion equation for this geometry to show that it is possible to derive the absorption and the reduced scattering coefficients of both layers if the relative reflectance is measured in the time domain at two distances and if the thickness of the first layer is known. Solutions of the diffusion equation for semi-infinite and homogeneous turbid media are also applied to fit the reflectance from the two-layered turbid media in the time and the frequency domains. It is found that the absorption coefficient of the second layer can be more precisely derived for matched than for mismatched boundary conditions. In the frequency domain, its determination is further improved if phase and modulation data are used instead of phase and steady-state reflectance data. Measurements of the time-resolved reflectance were performed on solid two-layered tissue phantoms that confirmed the theoretical results.     

In vivo 783-channel diffuse reflectance imaging system and its application

A fiber-based reflectance imaging system was constructed to produce in vivo absorption spectroscopic images of biological tissues with diffuse light in the cw domain. The principal part of this system is the 783-channel fiber probe, composed of 253 illumination fibers and 530 detection fibers distributed in a 20x20 mm square region. During illumination with the 253 illumination fibers, diffuse reflected lights are collected by the 530 detection fibers and recorded simultaneously as an image with an electron multiplying CCD camera for fast data acquisition. After signal acquisition, a diffuse reflectance image was reconstructed by applying the spectral normalization method we devised. To test the applicability of the spectral normalization, we conducted two phantom experiments with chicken breast tissue and white Delrin resin by using animal blood as an optical inhomogeneity. In the Delrin phantom experiment, we present images produced by two methods, spectral normalization and reference signal normalization, along with a comparison of the two. To show the feasibility of our system for biomedical applications, we took images of a human vein in vivo with the spectral normalization method.     

Nondestructive quantification of chemical and physical properties of fruits by time-resolved reflectance spectroscopy in the wavelength range 650-1000 nm

Time-resolved reflectance spectroscopy can be used to assess nondestructively the bulk (rather than the superficial) optical properties of highly diffusive media. A fully automated system for time-resolved reflectance spectroscopy was used to evaluate the absorption and the transport scattering spectra of fruits in the red and the near-infrared regions. In particular, data were collected in the range 650-1000 nm from three varieties of apples and from peaches, kiwifruits, and tomatoes. The absorption spectra were usually dominated by the water peak near 970 nm, whereas chlorophyll was detected at 675 nm. For all species the scattering decreased progressively with increasing wavelength. A best fit to water and chlorophyll absorption line shapes and to Mie theory permitted the estimation of water and chlorophyll content and the average size of scattering centers in the bulk of intact fruits.     

Compact tissue oximeter based on dual-wavelength multichannel time-resolved reflectance

We developed a compact dual-wavelength multichannel tissue oximeter based on the time-correlated single-photon counting (TCSPC) technique. The light sources are two pulsed diode lasers (output wavelengths of 672 and 818 nm, an average power of 1 mW, a pulse duration of 100 ps, and a pulse-repetition rate as high as 80 MHz). The time-resolved reflectance photons are detected by a multianode photomultiplier, and the output signals are redirected by a router to different memory blocks of the TCSPC personal computer board. The system's accuracy in determining the absorption microa and the reduced-scattering micros' coefficients and in reconstructing absorber concentrations in diffusive media was tested on phantoms. Preliminary in vivo tissue-oxygenation measurements were performed on healthy volunteers under different physiological conditions with a minimum acquisition time of 100 ms and an injected power of less than 100 microW.     

Noninvasive in vivo measurement of venous blood pH during exercise using near-infrared reflectance spectroscopy

Blood pH is an important indicator of anaerobic metabolism in exercising muscle. This paper demonstrates multivariate calibration techniques that can be used to produce a general pH model that can be applied to spectra from any new subject without significant prediction error. Tissue spectra (725 approximately 880 nm) were acquired through the skin overlying the flexor digitorum profundus muscle on the forearms of eight healthy subjects during repetitive hand-grip exercise and referenced to the pH of venous blood drawn from a catheter placed in a vein close to the muscle. Calibration models were developed using multi-subject partial least squares (PLS) and validated using subject-out cross-validation after the subject-to-subject spectral variations were corrected by mathematical preprocessing methods. A combination of standard normal variate (SNV) scaling and principal component analysis loading correction (PCALC) successfully removed most of the subject-to-subject variations and provided the most accurate prediction results.     

In vivo glucose measurement by surface-enhanced Raman spectroscopy

This paper presents the first in vivo application of surface-enhanced Raman scattering (SERS). SERS was used to obtain quantitative in vivo glucose measurements from an animal model. Silver film over nanosphere surfaces were functionalized with a two-component self-assembled monolayer, and subcutaneously implanted in a Sprague-Dawley rat such that the glucose concentration of the interstitial fluid could be measured by spectroscopically addressing the sensor through an optical window. The sensor had relatively low error (RMSEC = 7.46 mg/dL (0.41 mM) and RMSEP = 53.42 mg/dL (2.97 mM).     

Evaluation of physical properties of human hair by diffuse reflectance near-infrared spectroscopy

The objective of the present study is to develop a novel nondestructive, simple, and quick method to evaluate the friction, twist, and gloss of human hair based on near-infrared diffuse reflectance (NIR-DR) spectroscopy and chemometrics. NIR-DR spectra were measured for human hair, which was collected from eleven Japanese women (age 5-44 years), by use of an optical fiber probe. Partial least squares (PLS) regression has been applied to the NIR-DR spectra of human hair after mean centering (MC), standard normal variate (SNV), and first derivative (1d) or second derivative (2d) analysis to develop calibration models that predict the friction, twist, and gloss of human hair. We identified the most suitable wavenumber region for the evaluation of each physical property. Correlation coefficients and standard errors of calibration of the PLS calibration models for the friction, twist, and gloss of hair were calculated to be 0.96 and 0.023, 0.81 and 3.27, and 0.90 and 0.36, respectively. Thus, the calibration models have high accuracy.     

In vivo detection of dysplastic tissue by Raman spectroscopy

The detection of dysplasia and early cancer is important because of the improved survival rates associated with early treatment of cancer. Raman spectroscopy is sensitive to the changes in molecular composition and molecular conformation that occur in tissue during carcinogenesis, and recent developments in fiber-optic probe technology enable its application as an in vivo technique. In this study, the potential of Raman spectroscopy for in vivo classification of normal and dysplastic tissue was investigated. A rat model was used for this purpose, in which dysplasia in the epithelium of the palate was induced by topical application of the carcinogen 4-nitroquinoline 1-oxide. High quality in vivo spectra of normal and dysplastic rat palate tissue, obtained using signal integration times of 100 s were used to create tissue classification models based on multivariate statistical analysis methods. These were tested with an independent set of in vivo spectra, obtained using signal collection times of 10 s. The best performing model, in which signal variance due to signal contributions of the palatal bone was eliminated, was able to distinguish between normal tissue, low-grade dysplasia, and high-grade dysplasia/carcinoma in situ with a selectivity of 0.93 and a sensitivity of 0.78 for detecting low-grade dysplasia and a specificity of 1 and a sensitivity of 1 for detecting high-grade dysplasia/ carcinoma in situ.     

Noninvasive Raman spectroscopy of human tissue in vivo

We report the first transcutaneous Raman spectrum of human bone in vivo obtained at skin-safe laser illumination levels. The spectrum of thumb distal phalanx was obtained using spatially offset Raman spectroscopy (SORS), which provides chemically specific information on deep layers of human tissue, well beyond the reach of existing comparative approaches. The spectroscopy is based on collecting Raman spectra away from the point of laser illumination using concentric rings of optical fibers. As a generic analytical tool this approach paves the way for a range of uses including disease diagnosis, noninvasive probing of pharmaceutical products, biofilms, catalysts, paints, and in dermatological applications.     

In vivo fiber-optic confocal reflectance microscope with an injection-molded plastic miniature objective lens

For in vivo optical diagnostic technologies to be distributed to the developed and developing worlds, optical imaging systems must be constructed of inexpensive components. We present a fiber-optic confocal reflectance microscope with a cost-effective injection-molded plastic miniature objective lens for in vivo imaging of human tissues in near real time. The measured lateral resolution is less than 2.2 microm, and the measured axial resolution is 10 microm. Confocal images of ex vivo cervical tissue biopsies and in vivo human lip taken at 15 frames/s demonstrate the microscope's capability of imaging cell morphology and tissue architecture.     

High-information time-resolved Fourier transform spectroscopy at work

The first instrumental setup, to our knowledge, that is capable of recording in a few hours the time-resolved Fourier transform (TRFT) interferograms of gas-phase spectra that cover several thousands of inverse centimeters with spectral- and time-resolution limits that are equal, at best, to 2.5 x 10(-3) cm(-1) and 2 ns, respectively, is reported. It was developed on the stepping-mode Connes-type interferometer of the Laboratoire de Photophysique Moléculaire Université de Paris Sud. Also, for the first time, to our knowledge, these high-resolution TRFT spectra, illustrated with the Doppler-limited emission spectra of the N(2) transitions (B-A) and (B'-B) between 5500 and 11 000 cm(-1) and of the atomic Ar lines between 1800 and 4000 cm(-1), are recorded in the infrared spectral range. To obtain identical results that have the same signal-to-noise ratio, we should have increased the recording time of our unique previous high-information TRFT spectra by approximately 50,000. In other words, one hour is now long enough to obtain what would previously have required six years to record.     

Detection of bacteria by time-resolved laser-induced breakdown spectroscopy

A laser-induced breakdown spectroscopy technique for analyzing biological matter for the detection of biological hazards is investigated. Eight species were considered in our experiment: six bacteria and two pollens in pellet form. The experimental setup is described, then a cumulative intensity ratio is proposed as a quantitative criterion because of its linearity and reproducibility. Time-resolved laser-induced breakdown spectroscopy (TRELIBS) exhibits a good ability to differentiate among all these species, whatever the culture medium, the species or the strain. Thus we expect that TRELIBS will be a good candidate for a sensor of hazards either on surfaces or in ambient air.     

In vivo absorption spectroscopy of tumor sensitizers with femtosecond white light

A system based on a femtosecond white-light continuum and a streak camera was used for recordings of the in vivo absorption spectra of the tumor-seeking agent disulphonated aluminum phthalocyanine. Measurements for different drug doses were performed on tumor tissue (muscle-implanted adenocarcinoma) and normal muscle tissue in rats. It was found that the shape of the spectrum is tissue dependent. The peak of the absorption spectrum is blueshifted in tumor tissue as compared with the muscle. Thus the contrast in the drug-related absorption can be altered by up to a factor of 2 from the primary drug molecular-concentration contrast between normal muscle and tumor by the proper selection of the illumination wavelength.     

In vivo noninvasive measurement of blood glucose by near-infrared diffuse-reflectance spectroscopy

This paper reports in situ noninvasive blood glucose monitoring by use of near-infrared (NIR) diffuse-reflectance spectroscopy. The NIR spectra of the human forearm were measured in vivo by using a pair of source and detector optical fibers separated by a distance of 0.65 mm on the skin surface. This optical geometry enables the selective measurement of dermis tissue spectra due to the skin's optical properties and reduces the interference noise arising from the stratum corneum. Oral glucose intake experiments were performed with six subjects (including a single subject with type I diabetes) whose NIR skin spectra were measured at the forearm. Partial least-squares regression (PLSR) analysis was carried out and calibration equations were obtained with each subject individually. Without exception among the six subjects, the regression coefficient vectors of their calibration models were similar to each other and had a positive peak at around 1600 nm, corresponding to the characteristic absorption peak of glucose. This result indicates that there is every possibility of glucose detection in skin tissue using our measurement system. We also found that there was a good correlation between the optically predicted values and the directly measured values of blood samples with individual subjects. The potential of noninvasive blood glucose monitoring using our methodology was demonstrated by the present study.     

Specular reflection and diffuse reflectance spectroscopy of soils

Studies on the occurrence and effects of specular reflection in midinfrared spectra of soils have shown that distortions due to specular reflection occur for both organic (humic acid) and non-organic fractions (carbonates, silica, ashed fraction of soil). The results explain why the spectra of CaCO(3) in limed soils do not match published spectra and offer an explanation as to why the presence of inorganic C interferes with the development of calibrations for organic C. These results may also have implications for the use of mid-infrared spectra for quantitative and qualitative analysis of soils. For example, libraries of spectra collected by means other than diffuse reflectance would be largely useless for comparing mineral spectra to soil spectra. To obtain the best results with forages and grains, it is necessary to develop separate calibrations for different products, but this has not seemed to be a problem for diverse sets of soil samples with C contents of 0 to 5%. Mid-infrared calibrations have also appeared to be more robust than the corresponding near-infrared calibrations in that fewer outliers are found. However, the results discussed here indicate that at least for some soil types (e. g., large differences in mineralogy or C contents), separate calibrations may be necessary.     

In vivo determination of the molecular composition of artery wall by intravascular Raman spectroscopy

Atherosclerotic plaque vulnerability is suggested to be determined by its chemical composition. However, at present there are no in vivo techniques available that can adequately type atherosclerotic plaques in terms of chemical composition. Previous in vitro experiments have shown that Raman spectroscopy can provide such information in great detail. Here we present the results of in vitro and in vivo intravascular Raman spectroscopic experiments, in which dedicated, miniaturized fiber-optic probes were used to illuminate the blood vessel wall and to collect Raman scattered light. The results make clear that an important hurdle to clinical application of Raman spectroscopy in atherosclerosis has been overcome, namely, the ability to obtain in vivo intravascular Raman spectra of high quality. Of equal importance is the finding that the in vivo intravascular Raman signal obtained from a blood vessel is a simple summation of signal contributions of the blood vessel wall and of blood. It means that detailed information about the chemical composition of a blood vessel wall can be obtained by adapting a multiple least-squares fitting method, which was developed previously for the analysis of in vitro spectra, to account for signal contributions of blood.     

In vivo investigation of the immersion-liquid-induced human skin clearing dynamics

The results of an in vivo investigation of human skin clearing caused by the immersion-liquid-induced matching of the refractive indices are reported for the first time. It was established that subcutaneous injections of a glucose solution produce a significant long-term suppression of the light scattering in the skin dermis, which is an important factor ensuring an increase in efficacy of the optical tomography, photodynamic therapy, and photodestruction of deep objects.