Serial noninvasive photoacoustic imaging of neovascularization in tumor angiogenesis.

We present photoacoustic images of tumor neovascularization obtained over a 10-day period after subcutaneous inoculation of pancreatic tumor cells in a rat. The images were obtained from ultrasound generated by absorption in hemoglobin of short laser pulses at a wavelength of 1064 nm. The ultrasound signals were measured in reflection mode using a single scanning piezodetector, and images were reconstructed with a weighted delay-and-sum algorithm. Three-dimensional data visualize the development and quantify the extent of individual blood vessels around the growing tumor, blood concentration changes inside the tumor and growth in depth of the neovascularized region.

Monitoring cerebral perfusion using near-infrared spectroscopy and laser Doppler flowmetry.

This paper describes the simultaneous use of two, noninvasive, near-infrared techniques near-infrared spectroscopy (NIRS) and a continuous wave NIR laser Doppler flow system (LDF) to measure changes in the blood oxygenation, blood concentration and blood flow velocity in the brain. A piglet was used as animal model. A controlled change in the arterial CO2 pressure (PaCO2) was applied for achieving changes in the listed cerebrovascular parameters. The time courses of blood concentration parameters (NIRS) and RMS blood flow velocity (LDF) were found to correspond closely with those of carotid blood flow and arterial carbon dioxide pressure (PaCO2). This result shows the additional value of LDF when combined with NIRS, preferably in one instrument. Development of pulsed LDF for regional blood flow measurement is indicated.

Instrument-independent flux units for laser Doppler perfusion monitoring assessed in a multi-device study on the renal cortex.

To investigate the feasibility of instrument-independent perfusion units for laser Doppler flowmetry, a comparison was performed of two commercial fiberoptic laser Doppler perfusion monitors measuring the same flux situation for two different types of probes. In vivo measurements were performed on the cortex of pig's kidney, with an ultrasonic arterial flow meter as reference. The flow was mainly varied by internal arterial constriction using a balloon catheter. For each probe, instruments are compared in terms of the ratio of laser Doppler flux and arterial flow. For a given probe, the flux-to-flow ratios of the two instruments show a linear mutual relationship for a wide variety of arterial flows and laser Doppler fluxes. In vitro measurements were performed on an aqueous suspension of polystyrene microspheres. For the probe with interfiber distance 500 microm the ratio of the in vivo fluxes appears to agree within 16% to the value found in vitro, while for the 250-microm probe a difference of 28% was found. For a wide range of fluxes, the in vivo flux values of one instrument can be translated into flux values for the other instrument, in spite of the instrumental differences. This enables the user to render experimental results independent of the specific instrument, thus facilitating multi-center studies.

Photoacoustic imaging of brain perfusion on albino rats by using evans blue as contrast agent.

The visualization of the brain vascular system could be of great importance for studying its functionality and for diagnosing possible disorders. In this paper we report the use of photoacoustics for imaging brain perfusion on Albino rats in vivo and post mortem. The measurements on the animals were direct on the skin surface. The blood perfusion on skull cartilage was imaged and 2D slices were constructed by using a beamforming algorithm. From the images representation the Interactive Data Language (IDL, Research System Inc.) was used. We also investigated the possibility of using the Evans Blue dye as a substitute of blood for imaging brain structures in vitro. The breakdown of the dye under pulsed laser irradiation was studied and the energy under which this effect occurs was calculated for the wavelength of 532 nm.

Detection of photoacoustic transients originating from microstructures in optically diffuse media such as biological tissue.

The generation and detection of broadband photoacoustic (PA) transients may be used for on-axis monitoring or for imaging of optically different structures in the interior of diffuse bodies such as biological tissue. Various piezoelectric sensors are characterized and compared in terms of sensitivity, depth response, and directivity with respect to spherical broadband acoustic pulses. The influence on the sensor output of acoustic interference and refraction of the PA transients at the sample-sensor interface is discussed. Ring detectors are suitable for deep on-axis detection thanks to their strong directional sensitivity, and small disk sensors are most suited for 3-D imaging of microstructures such as the (micro)vascular system. Voltage and charge preamplification schemes are compared in terms of the signal-to-noise ratio (SNR). In all cases, the preamplifier noise turns out to be the limiting factor for the sensitivity. Based on experimental data, for several sensor types and optical wavelengths, the theoretical detectability of PA signals generated by blood-like absorbers in biological tissue is discussed.

Path-length distribution and path-length-resolved Doppler measurements of multiply scattered photons by use of low-coherence interferometry.

We report first results of measurements by low-coherence Doppler interferometry of the path-length distribution of photons undergoing multiple scattering in a highly turbid medium. We use a Mach-Zehnder interferometer with multimode graded-index fibers and a superluminescent diode as the light source. The path-length distribution is obtained by recording of the heterodyne fluctuations that arise from the Brownian motion of particles in an Intralipid suspension as a function of the optical path length. The experimental path-length distribution is in good agreement with predictions of Monte Carlo simulations. In the heterodyne spectrum, an increase of the mean Doppler frequency with path length is observed.

Image reconstruction for photoacoustic scanning of tissue structures.

Photoacoustic signal generation can be used for a new medical tomographic technique. This makes it possible to image optically different structures, such as the (micro)vascular system in tissues, by use of a transducer array for the detection of laser-generated wide-bandwidth ultrasound. A time-domain delay-and-sum focused beam-forming technique is used to locate the photoacoustic sources in the sample. To characterize the transducer response, simulations have been performed for a wide variety of parameter values and have been verified experimentally. With these data the weight factors for the spectrally and temporally filtered sensor signals are determined in order to optimize the signal-to-noise ratio of the beam former. The imaging algorithm is investigated by simulations and experiments. With this algorithm, for what is to our knowledge the first time, the three-dimensional photoacoustic imaging of complex optically absorbing structures located in a highly diffuse medium is demonstrated. When 200-mum-diameter hydrophone elements are used, the depth resolution is better than 20 mum, and the lateral resolution is better than 200 mum, independent of the depth for our range of imaging (to ~6 mm). Reduction of the transducer diameters and adaptation of the weight factors, at the cost of some increase of the noise level, will further improve the lateral resolution. The synthetic aperture algorithm used has been shown to be suitable for the new technique of photoacoustic tissue scanning.

Principles and practice of the laser-Doppler perfusion technique.

This paper reviews the development and use of laser-Doppler perfusion monitors and imagers over the past two decades. The enormous interest in microvascular blood perfusion coupled with the 'ease of use' of the technique has led to 1500+ publications citing its use. However, useful results can only be achieved with an understanding of the basic principles of the instrumentation and its application in the various clinical disciplines. The basic theoretical background is explored and definitions of blood perfusion and laser-Doppler perfusion are established. The calibration method is then described together with potential routes to standardisation. A guide to the limitations in application of the technique gives the user a clear indication of what can be achieved in new studies as well as possible inadequacy in some published investigations.

Doppler Monte Carlo simulations of light scattering in tissue to support laser-Doppler perfusion measurements.

Doppler Monte Carlo (DMC) simulations of the transport of light through turbid media, e.g., tissue, can be used to predict or to interpret measurements of the blood perfusion of tissue by laser-Doppler perfusion flowmetry. We describe the physical and mathematical background of Doppler Monte Carlo calculations, and present some comparisons with measurements, performed with experimental flow models, built to mimic skin tissue characteristics and for the calibration of perfusion instruments,which are important goals of the Biomed-I concerted action. The measurements deal with coherence effects.

Three-dimensional photoacoustic imaging of blood vessels in tissue.

We applied photoacoustics as a tissue tomography technique for the detection of blood concentrations, e.g., angiogenesis around tumors. We imaged blood vessels in highly scattering samples, using 532-nm light, to depths of ~1 cm . The samples were real tissue (chicken breast) or 10% dilutions of Intralipid-10%. The blood flowed through nylon capillaries. Polyvinylidene difluoride (PVdF) piezoelectric detectors were used in a surface-scanning mode. We demonstrate the sensitivity of the technique by photoacoustic detection of single red blood cells upon a glass plate. Lateral resolution is limited by the detector diameter (200 microm). The depth resolution is ~10 microm.

Feasibility of picosecond laser-Doppler flowmetry provides basis for time-resolved Doppler tomography of biological tissues.

A detectable signal is obtained from a laser Doppler flowmeter operating in the heterodyne mode with nano- and pico-second pulse laser sources. The ultrashort pulse probing may be useful for depth-dependent time-resolved laser Doppler velocity measurements of blood perfusion in biological tissues. © 1998 Society of Photo-Optical Instrumentation Engineers.

On refraction in Monte-Carlo simulations of light transport through biological tissues.

To obtain reliable results from Monte-Carlo simulations of light scattering experiments, a statistically accurate procedure for positioning the photons after refraction between two different scattering media is necessary. Two statistically equivalent algorithms for calculating the position of the photons immediately after crossing an interface are described and justified.

Probabilistic model of multiple light scattering based on rigorous computation of the first and the second moments of photon coordinates.

We consider a concise method based on recurrent relations that permit rigorous computing of the first and the second moments of the components of the vector locating a randomly walking photon in an infinite homogeneous light-scattering medium. On assumption that the components obey a three-dimensional Gaussian distribution a probability density for the photon locations at the Nth scattering event can readily be written down and the light-intensity distribution in the medium may be calculated. The results from theoretical analyses are compared with the solution of a light-diffusion equation and with results of Monte Carlo simulations and show a better fit with simulated data than the diffusion approximation.

Laser Doppler velocimetry and Monte Carlo simulations on models for blood perfusion in tissue.

Laser Doppler flow measurements and Monte Carlo simulations on small blood perfusion flow models at 780 nm are presented and compared. The dimensions of the optical sample volume are investigated as functions of the distance of the laser to the detector and as functions of the angle of penetration of the laser into the sample. The effects of homodyne and heterodyne scattering are investigated.

Laser Doppler blood flowmetry using two wavelengths: Monte Carlo simulations and measurements.

A new laser Doppler blood flowmeter for measuring skin perfusion is presented. The flowmeter consists of a probe that uses two different wavelengths and is able to measure at different depths. It may be used to distinguish the superficial microcirculation of the skin providing nutritional flow and the flow in deeper situated blood vessels (thermoregulatory flow). Measurements and Monte Carlo simulations of the Doppler signals for human skin are shown.

Fiber-coupled self-mixing diode-laser Doppler velocimeter: technical aspects and flow velocity profile disturbances in water and blood flows.

The effects of disturbances of the flow pattern in front of the fiber facet of a fiber-coupled self-mixing diode-laser Doppler velocimeter system are investigated. This was done by comparing measurements and calculations of the Doppler frequency spectrum with the expected values. The calculated Doppler spectrum was obtained from the calculation of light scattered (with or without Doppler shift) by the moving particles in front of the fiber facet. The velocity profile of the particles was calculated with a finite-element method. Measurements were done with water (with polystyrene spheres) and whole blood as the samples. Good agreement between measurements and calculations were obtained. The velocimeter was modeled as a five-mirror setup. The reflectivity of the fiber facet closest to the laser turns out to have the most influence on the sensitivity and stability of the laser. Direct reflection of unwanted light back into the laser cavity was avoided by placing a glass plate in front of the fiber. Design consi ations are presented.

A semiconductor laser used for direct measurement of the blood perfusion of tissue.

An instrument consisting merely of a semiconductor laser in its own housing was used to measure the blood perfusion in tissue. Use is made of the feedback of Doppler-scattered light to the photodiode in the laser housing. A recording perfusion of a finger under occlusion of blood flow in the arm is shown.

Condensed Monte Carlo simulations for the description of light transport.

A novel method, condensed Monte Carlo simulation, is presented that applies the results of a single Monte Carlo simulation for a given albedo micro(s)/(micro(alpha) & micro(s)) to obtaining results for other albedos; micro(s) and micro(alpha), are the scattering and absorption coefficients, respectively. The method requires only the storage of the number of interactions of each photon with the medium. The reflectance and transmittance of turbid slabs can thus be found from a limited number of condensed Monte Carlo simulations. We can use an inversion procedure to obtain the absorption and scattering coefficients from the total reflectance and total transmittance of slabs. Remitted photon densities from a semi-infinite medium as a function of the distance between the light source and the detector for all albedos can be found even from the results of a single condensed Monte Carlo simulation. The application of similarity rules may reduce further the number of Monte Carlo simulations that are needed to describe the influence of the distribution of scattering angles on the results.

Optical properties of human dermis in vitro and in vivo.

Condensed Monte Carlo simulation results have been used for calculating absorption and reduced scattering coefficients from the literature data on the measured total transmittance and total reflectance of samples of the human skin in vitro. The results of several measuring methods have been compared. We have also estimated the range for absorption coefficients and reduced scattering coefficients at 660 and 940 nm from measured intensities at the skin surface as a function of the distance from the location where the light enters the skin by using condensed Monte Carlo simulations for a homogeneous semi-infinite medium. The in vivo values for the absorption coefficients and the reduced scattering coefficients appear to be much smaller than the values from the in vitro measurements, that have been assumed until now. The discrepancies have been discussed in detail. Our in vivo results are in agreement with other in vivo measurements that are available in the literature.