Spatially distributed two-photon excitation fluorescence in scattering media: Experiments and time-resolved Monte Carlo simulations

We report on measurements and Monte Carlo analysis of the spatial distribution of light in two-photon excitation fluorescence (TPEF) microscopy operating through scattering media. The axial profile of TPEF produced by a high numerical aperture microscope objective focusing through a tissue-like optical phantom at depths more than five photon mean-scattering lengths is studied. The measured profile is quantitatively interpreted with time-resolved Monte Carlo simulations that take into account the spatio-temporal distribution of the photons within the turbid medium under a femtosecond excitation regime. It is shown that considering only the ballistic photons leads to an under-estimation of the actual out-of-focus distribution of the fluorescence, whereas it is over-estimated when the ballistic and scattered photons are treated alike. Comparison of the in-focus signal to the overall out-of-focus background clearly pointed out that the signal-to-background ratio of TPEF microscopy images benefits from a shortening of the excitation pulses. Moreover, in homogeneously labeled specimen, the background is shown to overcome the signal at imaging depths of about five to six photon mean-scattering paths.     

Analysis of light propagation in highly scattering media by path-length-assigned Monte Carlo simulations

Numerical analysis of optical propagation in highly scattering media is investigated when light is normally incident to the surface and re-emerges backward from the same point. This situation corresponds to practical light scattering setups, such as in optical coherence tomography. The simulation uses the path-length-assigned Monte Carlo method based on an ellipsoidal algorithm. The spatial distribution of the scattered light is determined and the dependence of its width and penetration depth on the path-length is found. The backscattered light is classified into three types, in which ballistic, snake, and diffuse photons are dominant.     

蒙特卡罗模拟光通过大气后的时间分布

采用蒙特卡罗方法模拟了光通过光学特性参数不同的大气后透射光时间分布.分析了大气的散射系数、吸收系数、不对称因子及折射率对透射光时间分布的影响.结果表明:透射光时间分布曲线存在两个峰,分别对应子弹光与漫射光.各光学参数界定了子弹光、蛇形光、漫射光的大小和时间范围,并从统计模拟的角度解释了折射率大的大气中难以成像的因为.     

Polarized optical coherence imaging in turbid media by use of a Zeeman laser

A method that uses a Zeeman laser in conjunction with a Glan-Thompson analyzer to image an object in a turbid medium is proposed. A heterodyne signal is generated only when the scattering photons are partially polarized, and the spatial coherence is not seriously degraded after the signal propagates in the turbid medium. A system combining polarization discrimination with optical coherence detection to image the object in a scattering medium is successfully demonstrated. The medium is a solution of polystyrene microspheres measuring 1.072 mum in diameter suspended in distilled water contained in a 10-mm-thick quartz cuvette. The advantages of this optical system, including better selectivity of the weak partially polarized scattering photons and better imaging ability in higher-scattering media, are discussed.     

Multiple Backscattering Effects in Optical Coherence Tomography Images of Layered Turbid Media

We study the influence of multiple backscattering on optical coherence tomography (OCT) images or pulsed-sounding images of layered turbid media. A method for calculating the signal of various orders of backscattering is proposed. The retarding effects of OCT-signal dropping are analyzed. The contrast attenuation of an image of a layered medium due to multiple backscattering is examined. The blurring effect of an image of the back boundary of a scattering layer, caused by a delay of multiply scattered photons compared with single-scattered ones, is studied theoretically and experimentally using an OCT device.     

Speckle diagnostics of multiple scattering media with the use of frequency-modulated laser radiation

A method for probing randomly inhomogeneous multiple scattering media with the use of frequency-modulated laser radiation is considered. The method is based on analysis of the dependence of the blinking index of time-averaged speckles formed upon scattering of the probing radiation in a medium on the frequency modulation depth of the probing radiation. In the case of a binary frequency modulation, the blinking index of the detected speckle-modulated radiation is determined by the cosine Fourier transform of the probability density of the optical path-length difference of partial components of the scattered field in the probed medium. The values of the probability density of the optical path-length difference reconstructed with the use of the proposed method from the measured blinking index of speckles for model scattering media (fluoroplastic layer and layer of TiO₂ particles on a glass substrate) are in a good agreement with the results of statistical simulation of the probing radiation transfer in multiple scattering media.     

Imaging Transparent Objects in a Turbid Medium Using a Femtosecond Optical Kerr Gate

A femtosecond optical Kerr gate time-gated ballistic imaging method is demonstrated to image a transparent object in a turbid medium. The shape features of the object are obtained by time-resolved selection of the ballistic photons with different optical path lengths, the thickness distribution of the object is mapped, and the maximum is less than 3.6%. This time-resolved ballistic imaging has potential applications in studying properties of the liquid core in the near field of the fuel spray.     

Polarization preservation in diffusive scattering from in vivo turbid biological media: effects of tissue optical absorption in the exact backscattering direction

There is considerable recent interest in using polarized light to investigate turbid biological media. Although tissue multiple scattering randomizes incident polarization states, there are circumstances when appreciable degree of polarization can be observed in diffusive scattering. In this study, we use polarization modulation and synchronous detection to examine in the exact backscattering direction the polarization properties of diffusely reflected visible light from hands of human volunteers of varying pigmentation levels. The surviving polarization fraction increases with increasing pigmentation, likely due to preferential loss of highly scattered, long-pathlength photons; this mechanism lowers the average pathlength traversed by the detected light and hence increases the measured polarization preservation. This behavior is contrasted with the overall diffuse reflectance intensity, whose magnitude decreases with increasing absorption. These experiments demonstrate the important influences of medium optical properties on the polarization characteristics of multiply scattered light, which must be further investigated to enable quantitative polarization evaluation of turbid media such as biological tissues.     

Laser optical feedback tomography

We describe a new method for imaging in three-dimensional turbid media, laser optical feedback tomography. This technique is based on the resonant sensitivity of a short-cavity laser to frequency-shifted optical feedback from ballistic photons retrodiffused from the medium. The advantage of the method is that the detector is the laser source itself, which provides optical amplification with self-aligned spatial and temporal coherent detection.     

Memory effect evaluation based on transmission matrix calculation

The memory effect is a type of auto correlation observed in linear systems, which is widely used to control scattered light through thin scattering layers.We show that there exists a strong correlation among the optimized phase distributions of adjacent focal points in focusing through scattering media.The numeric simulation and experiment indicate that within the memory effect, the phase difference between the two adjacent focal points shows an optical phase fringe pattern, and the closer the adjacent focal points are, the wider the fringe pattern will be, corresponding to the tilting of a plane wave phase added onto the acquired optical phase distribution at the focal point.This effect can be utilized for achieving optimal phase distributions of focal point scanning without optical phase evaluation via the experiment, which has great potential application in imaging through the scattering medium.     

Improved rejection of multiply scattered photons in confocal microscopy using dual-axes architecture

We perform Monte Carlo simulations to show that the dual-axes (DA) confocal architecture has superior rejection of multiply scattered photons in tissue to that of single axis. As a result, the DA configuration provides improved signal-to-noise ratio and dynamic range, and thus is sensitive to ballistic photons from deeper within tissue, features that are particularly useful for performing vertical cross-sectional reflectance images in tissue.     

Optical Computed Tomography Based on Extraction of Quasi-Straight forward Propagating Photons Using Sum-Frequency Generation Technique

Aiming at the realization of optical computed tomography (optical CT), a compact system comprised of two cw-lasers and a detector was proposed for the extraction of quasi-straightforward propagating photons from scattered light transmitting through a scattering medium. The extraction ability of the system based on the sum-frequency generation technique was investigated using a standard scattering medium of Intralipid-10% aqueous solution and was found to be 78 dB in dynamic range. The optical CT image of absorbers placed in the scattering medium was successfully obtained with high contrast using the proposed system.     

Bimodal temporal distribution of photons in ultrashort laser pulse passed through a turbid medium

The criteria were determined for simultaneous observation of scattered and unscattered (ballistic) peaks in temporal distribution of a narrow pulsed laser beam passed through a strongly scattering medium. Conditions were found for such an observation on the basis of non-stationary two-flux model for radiation transport. Calculation results permitted the definition of a compromise between the parameters of the initial laser pulse and the properties of a scattering medium. This provided the separate observation of various types of photons. The theoretical calculations were confirmed by experiments on recording pulse profiles of passed radiation of fs-laser in a water–milk solution.     

Acousto-optic laser optical feedback imaging

We present a photon noise and diffraction-limited imaging method combining an imaging laser and ultrasonic waves. The laser optical feedback imaging (LOFI) technique is an ultrasensitive imaging method for imaging objects through or embedded within a scattering medium. However, LOFI performances are dramatically limited by parasitic optical feedback occurring in the experimental setup. In this Letter, we have tagged the ballistic photons by an acousto-optic effect in order to filter the parasitic feedback effect and to reach the theoretical and ultimate sensitivity of the LOFI technique. We present the principle and the experimental setup of the acousto-optic laser optical feedback imaging technique, and we demonstrate the suppression of the parasitic feedback.     

偏振相移与相位共轭结合的散射光聚焦技术研究

散射介质对光的随机散射作用是制约其光学聚焦和成像的重要因素，光学相位共轭技术能够通过对散射光场共轭还原实现透过散射介质的光学聚焦和成像，其中散射光场相位的获取是共轭还原的核心。阐述了偏振相移的基本原理，将偏振相移与相位共轭技术相结合，设计了新的基于偏振相移的数字光学相位共轭系统。采用633 nm的HeNe激光器作为系统光源，毛玻璃作为散射介质开展散射光聚焦实验研究。实验结果表明:该装置能够成功实现透过散射介质的光学聚焦，其中聚焦点与背景光强的比值可达约400倍。     

Imaging in turbid media using quasi-ballistic photons

We study by means of experiments and Monte Carlo simulations, the scattering of light in random media, to determine the distance up to which photons travel along almost undeviated paths within a scattering medium, and are therefore capable of casting a shadow of an opaque inclusion embedded within the medium. Such photons are isolated by polarisation discrimination wherein the plane of linear polarisation of the input light is continuously rotated and the polarisation preserving component of the emerging light is extracted by means of a Fourier transform. This technique is a software implementation of lock-in detection. We find that images may be recovered to a depth far in excess of that predicted by the diffusion theory of photon propagation. To understand our experimental results, we perform Monte Carlo simulations to model the random walk behaviour of the multiply scattered photons. We present a new definition of a diffusing photon in terms of the memory of its initial direction of propagation, which we then quantify in terms of an angular correlation function. This redefinition yields the penetration depth of the polarisation preserving photons. Based on these results, we have formulated a model to understand shadow formation in a turbid medium, the predictions of which are in good agreement with our experimental results.     

基于光调制技术的混浊介质空间散射光信号检测方法研究

为提高被测混浊介质样品漫反射、漫透射和准直透射光信号的测量信噪比,首次将光调制技术运用于空间多点特征光信号的检测中。测量系统以调制盘转动频率对样品入射光强进行频率调制,然后对采集的多点光强数据通过离散傅里叶变换进行解调,提取调制频率点的谐波分量作为测量值。用该方法对浓度为8%,直径为0.7μm的聚苯乙烯小球悬浮液进行了实验测试,结果表明该方法能有效提高被测样品特征光信号的测量信噪比,为强噪声背景下混浊介质光学参数研究提供了新的途径和方法。     

Simulation of coherent properties of photons in scattering medium for optical coherence tomography

Monte Carlo model of optical coherence tomography is developed for simulation of photon transport in half infinite homogenous media. The procedure is accelerated by scaling the baseline data from standard Monte Carlo calculation in turbid media with arbitrary optical parameters. Gaussian beam is modeled by hyperboloid of one sheet for actual condition to obtain distribution of photons on sample surface. Depth dependence coherent signal and photons distribution are calculated in this way, which is important to reconstruction of optical parameters by inverse Monte Carlo. Numerical results have verified this method in turbid medium of different optical parameters with acceptable relative errors.     

Effect of absorption of multiply scattering media on the degree of residual polarization of backscattered light

The effect of absorption in a scattering medium on the degree of residual polarization of backscattered radiation is studied in the case of probing of multiply scattering media by a linearly polarized light. An approximate expression describing the dependence of the degree of residual linear polarization of the backscattered radiation on the optical characteristics of a multiply scattering medium is derived within the framework of the phenomenological approach, based on the concept of the distribution of the optical paths of partial components of the scattered optical field under the conditions of multiple scattering, and with the use of the ideas about the similarity of statistical moments of the multiply scattered optical fields. The cut-off of the partial components, characterized by a large value of the optical path, because of their absorption, results in a substantial increase of the degree of residual polarization for the bands of the selective absorption caused by the presence of chromophores in the scattering medium. The results of experiments with model scattering media (whole milk) and biological tissues (human skin in vivo) are presented.     

Polarization visualization of scattering media with CW laser radiation

The method of polarization visualization of a multiply scattering medium containing macroinhomogeneities based on analysis of polarization spatial distribution of a scattered linearly polarized light is discussed. The treatment is based on statistical properties of the effective optical path distribution of scattered field components. The influence of media scattering properties and the geometry of the experiment on the inhomogeneity image contrast obtained with use of polarization degree and of normalized scattered intensity of radiation as visualization parameters are discussed, as well as spatial resolution achieved in these both cases. Using the results of theoretical analysis and of the experimental model, the relationship between the shapes of spatial distributions of polarization degree and the intensity of the scattered light is considered as a function of the position of the visualized object (an absorbing half-plane immersed in a plane layer of the scattering medium). The opportunities for enhancing the quality of the images formed in this way are also discussed.