Laser light, when scattered from a moving object or fluid, fluctuates in intensity. These fluctuations can yield information about the velocity of the scatterers involved. Two approaches have been used over the past twenty years or so. The techniques of photon correlation spectroscopy and laser Doppler effectively measure the frequency shift that occurs when light is scattered from moving objects. Regarding the phenomenon as a time-varying speckle pattern, however, opens up several other measurement possibilities. This review paper describes some recent biomedical applications of these techniques.

The determination of the mean velocity of biological objects provides essential information about different biological systems: (1) motility measurements of microorganisms to test the water quality, (2) motility test of spermatozoa, and (3) blood flow measurements in capillaries. A description of these phenomena in terms of the speckle effect shows the correlation between the spectrum of the time dependence of the speckle pattern and the mean object velocity. No severe assumptions must be made. In the case of blood flow, one can differentiate between the blood flow and a superimposed movement of the surrounding tissue.

The theoretical and experimental investigations of the focused Gaussian beam (FGB) diffraction both in blood and lymph microvessels have been carried out. Speckle-interferometrical technique with the spectral analysis of scattered light intensity fluctuations has been applied for the investigation of lymph circulation in native microvessels. The measuring errors ofbioflow velocity has been analyzed. Alterations caused by the drug influence on lymph vessels have been studiecL Doppler method using FGB scattering is considered theoretically and experimentally. Keywords: focused Gaussian beam, diffraction. speckles, correlation function. lymph flow

We consider the transport of the electric field temporal autocorrelation in heterogeneous, fluctuating turbid media. Experiments are performed in strongly scattering media with spatially separated static and dynamic components, and low resolution `dynamical' images of such media are obtained using autocorrelation measurements of the emerging speckle fields taken along the sample surface. Our analysis, based on a diffusion approximation to the field correlation transport equation, reveals that the field correlation scatters from macroscopic dynamic heterogeneities within turbid media. Demonstrations using heterogeneous samples containing particles undergoing Brownian motion and shear flow are described.

Various approaches for the description of coherent beams scattering by tissues are discussed. Changes of object's scattering state due to the disease progress or drugs application are examined as the transition between different scattering modes. For the optically thin tissues demonstrating single-scattering regimes statistical and correlation analysis of the far-zone speckles produced by the broad or focused beam illumination allows to characterize and display scattering structures. Speckle technologies for tissue imaging and structure monitoring are designed.

The Hearing Organ has a coiled structure in which the basic geometrical arrangement of the different types of cells is repeated throughout the organ. Hearing organs of different mammalian species also utilize the same arrangement of cells. In order to understand the significance of this arrangement in processing the auditory stimuli it is essential to measure the cellular function in an intact organ of a living animal. The sensory cells in the intact inner ear of a living animal can be visualized in the basal turn of the cochlea through the round window membrane. To achieve this an imaging system is needed to detect the weak reflections from the sensory cells in the inner ear while rejecting strong reflections from structures below and above the region of interest, particularly the round window membrane itself. A confocal microscope/interferometer was developed and built to visualize the sensory cells and to measure their vibration in response to sound applied to the ear. The measuring system and some of its performance characteristics are described.

The application of laser speckle and speckle-interferometric methods to cardiovibrations measurements is presented. The homodyne and heterodyne measuring techniques that use the Michelson mterferometer are compared. As shown, the space-time projections of the mterferometer output signal can be useftil for analysis of biovibrations. New diffraction methods based on speckles dynamics with a small number of scatterers are suggested that allow pulse waves to be sensed and recorded without distortion. Keywords: speckles, interferometiy, cardiovibrations, pulse waves

The diffraction of a laser beam with a spatial amplitude-phase modulation in the form of regular interference pattern behind random phase object as biological tissue model is studied. The interference effects of speckle-fields formed in diffraction zone are considered. The concept of interference of mutually shifted partially developed identical speckle fields has been employed. The evolution of visibility of average intensity fringes formed in scattered field has been studied for the objects with different statistical parameters modeling various cell structures including objects with an oscillating correlation coefficient of phase inhomogeneities. The behavior of the speckle field interference induced by the focused spatially-modulated laser beam diffraction on the random phase objects has been studied. A new measuring technique of statistical parameters of transparent or reflecting random phase objects is discussed. The technique is based on the usage of the probing beam with regular interference pattern, focused on a moving object, and on measurement of contrast of average intensity fringes, formed in scattering dynamic speckle-modulated field. Biomedical applications of the developed modeling and measuring technique are discussed.

From a cw-laser beam transmitted to strongly scattering tissue most photons have been scattered and therefore a complicated unknown path in tissue and decoding of the optical tissue parameter from the intensity of the scattered photons is very difficult. But some of the transmitted photons could undergo the absorption and scattering. From its intensity the integral sum of the absorption and scattering coefficient can be calculated easily for its well known straight way. With a focused laser beam by scanning the sample in x, y and z-direction the optical parameters can be calculated with high lateral resolution. To distinguish between scattered and non-scattered photons, a Mach-Zehnder interferometer can be employed.

The qualitative analysis of human corneal endothelial cells patterns is evaluated with Fourier transform methods. The input images are gray level photomicrographs of in vivo human corneal endothelial cells obtained with a clinical specular microscope. The digital Fourier transform of the pattern of endothelial cells is obtained from the digitized gray level specular photomicrographs. The advantage of the Fourier transform analysis is the parallel processing of the Fourier transform and the potential development of a rapid system of analysis for large numbers of endothelial specular photomicrographs.

A new, real-time, flying slit confocal microscope, that has unique features and imaging characteristics for in vivo human ocular imaging is described. The use of scanning slit confocal microscopes has unique advantages over other instrumental systems based on pinhole-containing Nipkow disks (tandem-scanning confocal microscopes) for clinical in vivo confocal microscopy. The use of laser based confocal microscopy to investigate the metabolic state of biological tissues is described as a complementary technique to the reflected light imaging methods based on the flying slit confocal microscope.

The paper is devoted to experimental study of a new Laser Fourier Holographic Microscope with a digital image reconstruction stage. The ultrahigh contrast of the images of convenient biological tissues, as compared to an industrial high-performance Nikon compound microscope, is obtained and the reasons for that are discussed. A way for solution of a problem of speckle-noise for imaging systems is offered.

We have evaluated secondary laser induced retinal effects in non-human primates with a Rodenstock confocal scanning laser ophthalmoscope. A small eye animal model, the Garter snake, was employed to evaluate confocal numerical aperture effects in imaging laser retinal damage in small eyes vs. large eyes. Results demonstrate that the confocal image resolution in the Rhesus monkey eye is sufficient to differentiate deep retinal scar formation from retinal nerve fiber layer (NFL) damage and to estimate the depth of the NFL damage. The best comparison with histological depth was obtained for the snake retina, yielding a ratio close to 1:1 compared to 2:1 for the Rhesus. Resolution in the Garter snake allows imaging the photoreceptor matrix and therefore, evaluation of the interrelationship between the primary damage site (posterior retina), the photoreceptor matrix, and secondary sites in the anterior retina such as the NFL and the epiretinal vascular system. Alterations in both the retinal NFL and epiretinal blood flow rate were observed within several minutes post Argon laser exposure. Unique aspects of the snake eye such as high tissue transparency and inherently high contrast cellular structures, contribute to the confocal image quality. Such factors may be nearly comparable in primate eyes suggesting that depth of resolution can be improved by smaller confocal apertures and more sensitive signal processing techniques.

The theoretical model of coherent light scattering on enamel and dentin surface is described. The surfaces of hard tooth tissues are represented as 3D rough structures. The enamel components are assumed to form mosaic structure consisting of enamel prisms and interprism space elements. The possible tilt of enamel prisms is accounted. The dentin elements are represented as a set of shell-enclosed cylinders placed into bulk substance. The presence of peritubular dentin with refraction and absorption indices different to bulk dentin ones inside dentinal tubules is accounted. The results of computer simulation of laser radiation scattering on rough surfaces of enamel and dentin are represented. The dependencies of far-field axial intensity and FWHM of back-scattered light field on parameters of hard tooth tissue surfaces are calculated. The experimental set-up for examination of hard tooth tissues surfaces via parameters of back-scattered He-Ne laser radiation is designed. The investigation of laser radiation back-scattering on enamel and dentin surfaces is carried out. The experimental results are compared with theoretical predictions.

In the past interferometry has been applied in three main fields. Fringe interferometry has been used for the topographic measurement of surfaces. Interferometric length measurement techniques have been used for the measurement of distances, refractive indices, and wavelengths. In addition interferometric techniques have also been used in synthetic aperture imaging. Here we discuss various interferometric techniques which can be used in ophthalmology, describe the problems involved and present the results obtained so far.

Partial coherence tomography (or optical coherence tomography) uses signals obtained with partial interferometry techniques to synthesize tomographic images. Being still under evolution this technique has already been successfully applied in the clinic. We discuss the underlying principles and present some results obtained at the fundus of the eye.

A broadband interferometer is used to acquire scattered light as a function of depth in biological media. The `tissue-light-signature' that is obtained by this depth scan can be correlated with the computer simulated light distributions for well defined tissue parameters, and wavelengths of specific interest. In theory, the collimated irradiation of heart tissue, by low coherence light will generate a statistically significant different light signature for respective myocardial tissues, and pathological tissue conditions. Interferometric axial scanning of in vitro myocardial tissues confirmed the statistically significant difference between normal, coagulated myocardium, and aneurysm at the 790 nm wavelength. The scanning depth however is presented limited by the intensity of the illumination and the choice of detection scheme. Identification of the local optical characteristics as a function of depth directly underneath the target zone will provide discrimination between healthy and pathological conditions in addition to real time assessment of laser dosimetry. Theoretically the scanning depth is limited to a maximum of 4 mm. The beam profile of the irradiation source significantly affects the ability to distinguish between certain tissues. Broadband interferometric axial tissue scanning, will provide a tool for an accurate light energy delivery guided by the desired outcome, while being able to verify the appropriate target location, in real time.

When performing direct-contact laser-Doppler flowmetry on experimental flow models, the power spectra of the detector signal can be obtained by homodyne or by heterodyne detection. Especially with uniformly moving probe particles coherence effects are observed, leading to changes in the width of the power spectrum. Due to destructive interference effects at the detector surface, in the measured homodyne spectra the contribution of the relatively high Doppler frequencies is suppressed compared with that of lower frequencies. This results in spectra which are narrower than expected theoretically. This effect allows us to investigate the importance of the relative amount of coherent areas at the surface of the detector.

Optical metrology based on the principle of interference can be applied as a testing tool in biomedical research. Interferograms in form of fringe patterns can be produced in two-beams interferometers, holographic or speckle interferometers, in setups realizing moire techniques or in deflectometers. By analyzing of the fringe pattern images, information about the shape or mechanical behavior of the object under study can be retrieved. Here, some of the techniques for creating fringe pattern images were presented along with methods of analysis. Possible applications of interferometric methods, especially in the field of experimental orthopedics, endoscopy and ophthalmology will be pointed out.