We have already demonstrated the potentiality of interferometry to perform timeresolved
measurements of the light scattered by a tissue: the fluctuations of the speckle
pattern, linked to a wavelength-modulation of the source, are registered, and the time-resolved
average intensity can be numerically obtained from these data[1]. The competitive results were
obtained with a simple photodiode as detector[2].
Such a method can be cheaper and more accessible for biomedical applications than
direct time-resolved methods, but it is not its unique advantage: this method allows to perform
Diffusing Wave Spectroscopy (DWS) with selected photon pathlengths[3,4]; for instance, we
have shown that we can improve the spatial resolution in transillumination imaging of a
dynamic heterogeneity through the selection of short photon transit times[4]. Therefore such a
method can offer interesting applications, for example in mammography.
A way to improve the signal to noise ratio of this method can consist in multiplying
the number of detectors. That's the reason why we decide to consider the use of a high speed
camera, that can reach a rate of 1000 frames per second. We will present the first results
obtained with this new system. The performance will be discussed, and compared to our
previous setup.
Diffusing Wave Spectroscopy (DWS) consists in the measurement of temporal correlation of the electromagnetic field in the diffusion regime, allowing a scan of dynamical properties deep inside a medium. DWS is of special interest in biomedical optics, as it is sensitive to blood circulation in capillaries inside the tissue.
However one main difficulty of this technique concerns data extraction which implies to perform an inverse problem taking into account the geometry and the optical coefficients of the medium. The use of time-resolved detection has been proved to be an efficient tool to discriminate the DWS information, but the photon path lengths were up to now limited to a few tens mean free paths.
In order to perform time-resolved DWS for much longer photon paths, we used a new method, based on the use of an interferometer and a wavelength modulated source. We have already demonstrated that this method, in addition to its lower cost, was very efficient to perform time-resolved measurements of the light scattered by a thick scattering medium. We will show in this poster some measurements performed by transillumination through a thick medium (4cm), opening the possibility of Time-Resolved DWS measurements in the human breast.
KEYWORDS: Tissue optics, Sensors, Reflectivity, Picosecond phenomena, Streak cameras, Scattering, Optical testing, In vivo imaging, Dielectric filters, Monte Carlo methods
We develop a new sensor for the local in vivo measurement of optical coefficients near the surface of a tissue. To be less sensitive to the heterogeneous surface of the sample, we decided to perform space and time-resolved measurements. The sensor is a bundle of fibres. The
excitation light is generated by a mode-locked Ti-Sa laser at 800nm and filtered by a 1.5nm bandwidth dielectric filter in order to limit group velocity dispersion in the monomode excitation fibre. The reflectance light is collected by gradient index fibres at 250μm and 1.3 mm from the source. The detection is performed with a Hamamatsu M5675 synchroscan streak camera. The whole system allows a time resolution of about 5ps. We made comparisons between time and space resolved Monte-Carlo numerical simulations and in vitro experimental data obtained with unskimmed UHT milk which is a known reference medium. The system does not rely on the absolute value of the reflected light intensity nor depend on the intensity ratio between different fibres since the distance between the medium and the fibres as well as the fibres tip cleanness cannot be guaranteed in vivo. As a consequence we use global characteristic of the time resolved curves such as the FWHM and their evolution with the distance from the source. The good agreement between the simulations and the experimental data lets us envisage to use numerically pre-computed tables for a real time determination of the local scattering mean free path and the anisotropy factor . We soon will be able to perform
measurements with biological tissues, in vitro in a first time and in vivo in a second time.
A clinical study of the evolution of the space resolved near infrared reflectance with the subcutaneous layers thickness, at rest, on the vastus lateralis, is used to determine convenient sets of muscle and fat optical coefficients at 850 nm and 780 nm. These coefficients are then used to establish pre-calculated 3 layers MC simulations tables of the reflectance as a function of the fat thickness. Real time inversion during effort is then performed to determine the absolute muscle oxygenation.
A clinical study of the evolution of the space resolved reflectance with the subcutaneous layers thickness, at rest, on the vastus lateralis, is used to determine convenient sets of muscle and fat optical coefficients at 850 nm. The results are compared to the data available in the literature. When the same study will be completed at 780 nm, these coefficients will be used in 3 layers MC simulations to establish pre-calculated tables of the reflectance as a function of the fat thickness and of the muscle absorption coefficient. The goal of this study is eventually to perform real time inversion during effort in order to determine the muscle oxygenation with this very simple technique (part 2).
We explore in this paper a new method to perform time-resolved measurements of the diffuse light transmitted through a thick turbid medium. This method is based on the analysis of the speckle fluctuations due a wavelength modulated source. A time resolution of about 50 ps is already achieved, and we expect to improve this result soon. This method could allow the design of low cost setups to perform such measurements.
We have already demonstrated that some optical coefficients of turbid media ((mu) a, (mu) s') can be derived from time and space resolved backscattered light measurements in the case of semi-infinite geometry, or multi-layered media, using the experimental results obtained far enough from the illumination area, that is where and when the diffusion approximation is valid. Our purpose is now to determine if this type of measurements can be used to explore the different moments of the scattering phase function. The same experimental set-up consisting of a titanium-sapphire pulsed laser and a streak camera was therefore used to measure the reflectance at earlier times and closer to the light source. Experiments were performed with aqueous solutions of calibrated latex microspheres. Various values of the spheres diameter were alternately used in order to vary the anisotropy factor g. Comparison with Monte-Carlo simulations were achieved for the resolution of the inverse problem. The influence of the shape of the phase function was also investigated.
Our purpose is to develop an optical technique for in-vivo and non-invasive diagnosis using backscattered light measurements. We have already demonstrated that optical coefficients of turbid media ((mu) a, (mu) s) can be derived from time and space-resolved reflectance in the case of semi-infinite geometry. This procedure was then applied to the investigation of multi-layered media: the upper layer was an aqueous solution of calibrated latex microspheres in water and the lower layer of the sample was a solid phantom. Two different types of phantoms were used. In the first set of experiments, we used an absorbing medium for under layer. In the second case, the lower layer was an absorbing and scattering phantom. Comparison with Monte-Carlo simulations were achieved for the resolution of the inverse problem.
Time and space resolved measurements of diffuse backscattered light form turbid media were performed to determinate the optical coefficients of biological tissues. The reflectance maps registered on a streak camera were analyzed with two different methods based on the diffusion approximation. These fast and practical methods were initially tested on suspensions of latex spheres in water to compare the experimental results with the theoretical coefficients derived from Mie theory. Measurements were then achieved on biological tissues.
The aim of this study was to perform a preliminary evaluation of the diagnostic potential of laser induced autofluorescence spectroscopy (LIAFS) for urothelial tumors using fluorescence intensity ratios at different wavelengths. After testing three laser excitation wavelengths (308, 337, and 480 nm) in normal and malignant bladder cell lines, 308 nm appeared to be the most promising wavelength since two fluorescence bands were observed at 360 and 440 nm; these were attributed to tryptophan (Trp) and reduced nicotinamide adenine dinucleotide (NADH) respectively. This study was then performed on freshly removed normal bladder and bladder tumor specimens exclusively using the 308-nm excitation wavelength. The tumor spectra, regardless of stage and grade, were very similar to the malignant cell spectra. However, a marked reduction of overall
intensity was observed for carcinoma in situ (CIS). Normal bladder mucosa exhibited a shift of the first
fluorescence band to 380 nm, indicating an overlap of Trp urothelial cell emission and collagen fluorescence
derived from the lamina propria. The intensity of the NADH emission band was markedly reduced in tumor
tissues compared with normal mucosa, which could indicate different redox conditions in urothelial tumors. A fluorescence intensity ratio at 360 and 440 nm can accurately discriminate normal or inflammatory mucosa from all bladder tumors, including CIS. These findings support the use of LIAFS as a new diagnostic technique for occult urothelial tumors.
We have designed a program using laser induced autofluorescence spectroscopy as a possible way to characterize urothelial tumors of the bladder. The autofluorescence spectra were compared between normal, suspicious and tumor areas of human bladder. Three different pulsed laser wavelengths were used for excitation: 308 nm (excimer), 337 nm (nitrogen) and 480 nm (dye laser). Excitation light was delivered by a specially devised multifiber catheter introduced through the working channel of a regular cystoscope under saline irrigation. The fluorescence light was focused into an optical multichannel analyzer detection system. The data was evaluated in 25 patients immediately before resection of a bladder tumor. Spectroscopic results were compared with histopathology. Upon 337 nm and 480 nm excitations, the overall intensity of the fluorescence spectra from bladder tumors was clearly reduced in comparison with normal urothelium, regardless of the stage and the grade of the tumor. upon 308 nm excitation, the shape of tumor fluorescence spectra, including carcinoma in situ, differed drastically from that of normal tissue. In this case, no absolute intensity measurements are needed and clear diagnosis can be achieved from fluorescence intensity ratio (360/440 nm). This spectroscopic study could be particularly useful for the design of a simplified autofluorescence imaging device for real-time routine detection of occult urothelial neoplastic lesions.
The aim of this paper was to demonstrate the feasibility of routine real time clinical detection of occult urothelial precancerous and cancerous lesions by laser induced autofluorescence (LIAF), that is with no previous instillation of exogenous fluorescent marker. Three different pulsed laser wavelengths were alternately used for excitation: 480 nm (dye laser), 337 nm (nitrogen laser), and 308 nm (XeCl excimer laser). A clinical endoscopic study was performed on 23 patients immediately before transurethral resection of bladder tumor. Spectroscopic results were compared with histological analysis. For 480 nm and 337 nm excitation a single fluorescence broad band was obtained in any case, but for tumors the overall intensity was significantly reduced compared to normal mucosa. For 308 nm excitation, two main broad bands were observed. In the case of neoplasic lesions (including carcinoma in situ), the intensity ratio [I(360 nm)/I(440 nm)] was always greater than 2, but for normal or inflammatory areas this ratio was less than 2. A clear diagnosis could then be achieved for 308 nm excitation without the need of absolute intensity measurements. We get no false positive at this wavelength. XeCl LIAF spectroscopy is therefore a promising technique for the detection of urothelial precancerous lesions and could be used to perform an 'optical biopsy' in a routine mode with real time results.
A collimated light beam propagating through a turbid medium can be strongly attenuated by both absorption and scattering. It is usually assumed that, in first approximation, the attenuation depends linearly on the thickness of the scattering medium and on the scatterers concentration. This law, based on the assumption that the scatterers are uncorrelated, is in good agreement with experimental measurements when the particles density is low. However, in a dense distribution of particles, it has been shown that corrections need to be introduced. In this paper, we relate some experimental results concerning coherent propagation in latex spheres suspensions in water. These results are obtained by the use of polarization analysis and show the deviation of the attenuation from the linear law in spheres concentration. A good agreement is obtained with the Keller propagation theory in dense media.
Recent biomedical optics experiments, imaging or quantitative measurements of chemical compounds for example, are more and more sensitive to the optical characteristics of biological tissues. Artificial scattering media are used in the laboratories in order to work in reproducible, stable and well known samples. The most difficult part of the work is to obtain an adequate phase function, since the scattering and absorption coefficients can be adjusted by an appropriate concentration of the scattering particles and the addition of an absorbing dye. The most forward way to create phantoms is to use scattering spheres of equal size. If the sphere diameter which gives the desired mean cosine of the single scattering angle is not available, similarity relations may provide the necessary adjustments. However, as we show in this paper, these similarity relations may sometimes be very inaccurate and, moreover, the Mie phase function of a sphere does not match a real tissue phase function. Arridge et al, Firbank et al have suggested that a better solution would be to use a distribution of different size scattering particles in order to imitate the whole phase function, but the determination of a mixture of spheres with adequate sizes and concentrations is a difficult mathematical problem. The goal of this paper is to solve this problem. It is first shown that the extreme complexity of real biological samples can be very simply simulated by a mixture of spheres with a fractal diameter distribution. Then some simple rules, based on the knowledge of this fractal distribution, are given in order to obtain a realistic phase function with a limited number of spheres diameters.
Laser induced fluorescence has often been used as a diagnostic method. Unfortunately the fluorescence signal is modified during the photons migration towards the detector. The purpose of this study is to determine the alterations of the laser induced fluorescence spectra in white matter of adult brain due to the spectral variations of the optical coefficients (mu) a((lambda) ), (mu) s((lambda) ) and of the mean cosine of the scattering angle g((lambda) ).
Three-hundred-eight nm laser-induced autofluorescence spectra of the normal human brain, astrocytoma grade IV and glioblastoma grade IV specimens, have been recorded in vitro two hours after surgical resection. Typical fluorescence spectra for normal (N) and malignant (M) tissue show 4 maxima at about 352, 362, 383, and 460 nm. These spectra are analyzed in detail. Subtle differences in normalized spectra of N and M tissues appear to be large enough for diagnosis. Several criteria such as maxima and minima absolute intensity and intensity ratios at typical wavelengths are computed and used to classify the tissue. This preliminary study shows that fluorescence spectroscopy with 308 nm UV excitation could be a valid technique for discriminating tumor types. However, it should be noted that these measurements are made in vitro. Living tissues may have different spectral characteristics, therefore future in vivo investigations must be performed.
We discuss the validity and the precision of scaling principles which can be used to transform problems of radiative transfer in media that scatter anisotropically (g does not equal 0, g being the mean cosine of the scattering angle) to equivalent isotropic problems (g equals 0). A Monte Carlo model is used to calculate the reflection (R) and the transmission (T) factors for a collimated incident beam on an anisotropically scattering media, for a wide range of absorption ((kappa) ) and scattering ((sigma) ) coefficients per unit length. The results (R,T) obtained for anisotropic scatterers (g does not equal 0, (kappa) , (sigma) ) are systematically compared to those (R', T') obtained in the case of the equivalent isotropic configuration (g' equals 0, (kappa) ', (sigma) ') as deduced from the similarity relations.
The design of a new semi-analytical Monte Carlo simulation is discussed in detail in this paper. This model uses two stages. In the first stage, the simulation itself, the contribution of each scattering event to the total reflectance and transmittance is evaluated. Thus the photon energy decreases more rapidly during its random walk and fewer steps are required to obtain a given accuracy. The reduced number of necessary steps makes it possible to store all events positions and energies. In the second stage, the results of the first stage can be used to calculate analytically any desired result. Examples are given for scattering slabs of isotropic or anisotropic scatterers when collimated beam incidence is used. Reflections at the boundaries are taken into account. The results obtained with this new method and classical Monte Carlo methods are identical. However, the convergence of our new model is much better and, because of the separation in two stages, any quantity related to the problem can be easily calculated afterwards without recomputing the simulation.
In this theoretical paper we study the alterations of the laser induced fluorescence spectra as a function of the spectral variations of the absorption factor μa(λ), the scattering factor μs(λ) and of the mean cosine of the scattering angle g(λ) of the medium. The calculations of the signal received by the detector are based on a Monte Carlo simulation. All media are homogeneous, with infinite length and width, depth is assumed to be infinite. The total detected fluorescence signal S(λ) may be described by: S(λ)= F(λe,λ) . M(μa(λe),μs(λe),g(λe),μa(λ),μs(λ),g(λ)) where F is proportional to the original fluorescence spectrum and M a function of the optical and geometrical properties of the medium. In order to show the importance of the scattering, absorption and anisotropy spectral dependence, we have computed M for ideal experiments. Each coefficient μa, μs and g is alternately considered to be wavelength dependent while the others are considered to be constant. Consequently, each calculation provides a different distorted transfer function M(λe,λ) showing the influence of the spectral dependence of each coefficient. This signal is also different if the wavelength of the excitation light is changed.
KEYWORDS: Brain, Tissues, Laser tissue interaction, Excimer lasers, Gas lasers, Fiber optics, In vivo imaging, Control systems, Laser therapeutics, Magnetic resonance imaging
Deep brain tissue lesions produced by XeCl excimer laser were studied in 14 rats. Fiberoptic was stereotactically implanted and the lesion was produced by one pulse. MRI and histological controls were performed at various dates following the lesion. The findings are relevant to other lesion procedures. Some XeCl laser properties seem useful in stereotactical procedures.
We present the results of 12 XeC1 laser coronary artery endarterectomies performed in 10 patients during
CABG surgery. The results are very encouraging and led us to the developement of a new and more efficient laser
catheter &livery system which is described in the second part of this paper.
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