We present the results of the first quantitative multimodal confocal imaging study of methylene blue (MB)-stained cancer and normal human renal cells obtained from fine needle aspiration (FNA) biopsies. Fluorescence emission images provided morphological assessment, and fluorescence polarization (Fpol) images yielded quantitative characterization of each cell in the investigated samples. FNA specimens are obtained from discarded malignant and normal renal specimens following surgery. Prior to imaging, the cells are stained in aqueous MB solution. Our results demonstrate that all the specimens investigated are heterogeneous in terms of size and exhibited Fpol. Cancerous specimens predominantly contain cells of larger size that exhibit higher Fpol as compared to normal specimens. Imaging results correlated well with clinical assessment of the samples. Our results suggest that morphological assessment using fluorescence emission imaging and quantitative information provided by Fpol imaging may be valuable in determining the presence or absence of renal cancer cells in FNA specimens.
Background and Objective: Extraoral photobiomodulation therapy (PBM Therapy) is a novel treatment for the prevention of oral mucositis, a painful side effect of myeloablative chemotherapy, and there are no standardized dosimetry protocols for this procedure. We used Monte Carlo modeling to determine optimal parameters for a safe and efficacious treatment. The objective of this work was to experimentally validate the results of Monte Carlo dose modeling of extraoral PBM Therapy by conducting a pilot validation study.
Methods: Light penetration through the right cheek of four volunteers with skin types I-VI was measured. A 69-LED array with an area of 31.2 cm2 was applied to the external cheek delivering 26 mW/cm2 at 850 nm to the surface. Power density at the intraoral mucosa was recorded under a controlled pressure of 18gm/cm2.
To obtain morphological information, we acquired T1 weighted MRI images of the volunteers' heads and measured the thickness of the skin, fat, and muscle layers of the cheek of each volunteer. These anatomical data together with the optical properties from the literature were used to simulate light propagation through the right cheek.
Results and Conclusions: Our study revealed that experimental and simulation results were in good agreement for all 4 subjects. The difference between the mean values of the measured fluence rates was within 16% from the respective fluence rates obtained using Monte Carlo simulations. We confirmed that there was no temperature increase due to illumination. Monte Carlo modeling is a robust and reliable method for PBM Therapy light dosimetry.
Background and Objective: Intraoral photobiomodulation therapy (PBMT) is effective at preventing oral mucositis (OM). Extraoral PBMT is a novel approach with distinct advantages over intraoral PBMT, but no evidence-based treatment protocol has been proposed. The goal of this contribution was to develop a practical and effective treatment protocol for extraoral PBMT to prevent OM.
Methods: Extraoral PBMT was modeled using Monte Carlo simulations with parameters established in intraoral PBMT: 50 mW/cm2 (1 minute treatment time), at either 660 nm, used in superficial PBMT, or 850 nm, used in deeper PBMT. The effective therapeutic dose of ~ 2.0 J/cm2 was assumed to be identical for intraoral and extraoral PBMT. The results prompted a review of the literature regarding PBMT parameters for deep tissue treatment, as well as the mechanism of PBMT.
Summary: Per our Monte Carlo simulations, the treatment parameters established for intraoral PBMT are not appropriate for extraoral delivery. Visible light (660 nm) showed poor penetration, producing an insufficient dose at the oral mucosa. The infrared light (850 nm) penetrated deeper. However, at 50 mW/cm2 and 1 minute treatment time, the dose was still less than targeted 2 J/cm2. Therefore, based on the optical properties of tissues, anatomy of the target population, and established protocols for deeper PBMT, we propose a wavelength of 850 nm, at 399 mW/cm2, and exposure time of 1-11 minutes, depending on the treatment site, to achieve the therapeutic dose of ~2 J/cm2 at the depth of the oral mucosa for preventing OM using extraoral PBMT.
The objective of this study was to investigate the feasibility of using optical polarization imaging (OPI) for the preoperative delineation of nonmelanoma skin cancer (NMSC) margins. OPI has been previously used for monitoring dermal collagen changes in healthy volunteers in vivo. Since the structure of normal collagen is disrupted in NMSCs, OPI should be capable of delineating skin cancer margins by identifying the disrupted collagen network.
Patients with biopsy confirmed NMSCs were recruited under an IRB approved protocol. Prior to imaging, the lesion area was cleaned with alcohol and the intended boundaries of the excision were marked by the surgeon, who was blinded from the imaging results. The imager used narrow band linearly polarized light centered at 440 nm and 640 nm. Cross-polarized reflectance images acquired at 440 nm and 640 nm visualized dermal collagen and the blue marker used by the surgeon to outline the putative lesion boundaries, respectively. The imager provided a 4 cm2 field of view, 200-700µm imaging depth and the light power density at the skin surface of 0.38mW/cm2. Following imaging, routine Mohs procedure was performed using the original markings of the surgeon. After the surgery, images acquired by OPI were compared with the Mohs maps created based on histopathological analysis.
Our results indicate that OPI accurately predicts subclinical extension of NMSCs beyond visibly-involved margins in the majority of cases. Therefore, OPI holds the potential to reduce the total number of required surgical stages, possibly minimizing the unnecessary removal of normal tissue.
Background: Oral mucositis (OM) often occurs after myeloablative hematopoietic cell transplantation. We explore extra-oral photobiomodulation therapy (PBMT) for the prevention of OM in children. Objectives: Our objective was to use modeling of PBMT to determine optimal treatment sites and parameters for a safe and efficacious treatment. Methods: MRI images were analyzed to obtain morphological information on the extent of tissues along the six trajectories passing through the cheek; lips; submandibular and submental regions; neck, transversely and anteroposteriorly. For each treatment site we performed 18 simulations using morphological information from 18 subjects with ages between 5 and 20 years. The simulation technique uses Monte Carlo method to calculate light distribution and finite difference method to solve the heat transfer equation. Our model accounts for the geometry of emitter, optical, thermal, and morphological tissue properties, as well as possible changes of the optical properties during the therapy. We have used a layered tissue model. The optical and thermal properties of tissues were assigned to each pixel individually using published data on the properties of relevant tissues. We evaluated spatially resolved fluence rate, absorbed power, temperature increase and thermal damage. Results and Conclusions: At 850nm and 399mW/cm2, the median dose transmitted ranged from 0.18–2.4J/cm2. As presence of blood hinders light penetration, treatment sites should be chosen to avoid major blood vessels. There was no temperature increase and damage to the tissue. Our results reveal that extra-oral PBMT is safe and shows promise for the prevention of OM in children.
Background: Oral mucositis (OM) is a painful consequence of myeloablative hematopoietic cell transplantation (HCT).
Extraorally delivered photobiomodulation therapy (PBMT) is a promising novel intervention for the prevention of OM
in children. Objectives: With funding from an NIDCR R34 planning grant, the objectives of this study are 1) to model
the dosimetry of external PBMT and the optimal device parameters for the planned clinical trial, and 2) to plan and
design a placebo-controlled Phase 2 multicenter clinical trial to determine whether extraorally delivered PBMT can
reduce the duration of severe OM in children, with intent for implementation under subsequent U01 funding. Methods:
External PBMT dosimetry will be evaluated using pediatric head and neck MRI studies to obtain serial measurements of
different tissues that will then be used to develop a sophisticated computational model. We plan to conduct a placebocontrolled
Phase 2 multicenter clinical trial in which patients 4 to 21 years of age will be randomized 1:1:1 to receive
external PBMT dose 1x, external PBMT dose 2x, or sham PBMT starting from conditioning, daily until day +20 post-
HCT. Significance: Extraorally delivered PBMT is a feasible, potentially efficacious intervention that could improve the
quality of life for all children undergoing myeloablative HCT. The planned Phase 2 study, based on rigorous dose
modeling and with detailed attention to uniform delivery of therapy and OM assessments, will provide critical efficacy
data and the potential basis for a subsequent definitive Phase 3 trial. Grant Support: NIDCR R34 DE025908-01
Contrast agents have shown to be useful in the detection of cancers. The goal of this study was to compare enhancement of brain cancer contrast using reflectance and fluorescence confocal imaging of two fluorophores, methylene blue (MB) and demeclocycline (DMN). MB absorbs light in the red spectral range and fluoresces in the near-infrared. It is safe for in vivo staining of human skin and breast tissue. However, its safety for staining human brain is questionable. Thus, DMN, which absorbs light in the violet spectral range and fluoresces between 470 and 570 nm, could provide a safer alternative to MB. Fresh human gliomas, obtained from surgeries, were cut in half and stained with aqueous solutions of MB and DMN, respectively. Stained tissues were imaged using multimodal confocal microscopy. Resulting reflectance and fluorescence optical images were compared with hematoxylin and eosin histopathology, processed from each imaged tissue. Results indicate that images of tissues stained with either stain exhibit comparable contrast and resolution of morphological detail. Further studies are required to establish the safety and efficacy of these contrast agents for use in human brain.
The goal of the study was to evaluate wide-field and high-resolution multimodal optical imaging, including polarization, reflectance, and fluorescence for the intraoperative detection of breast cancer. Lumpectomy specimens were stained with 0.05 mg / ml aqueous solution of methylene blue (MB) and imaged. Wide-field reflectance images were acquired between 390 and 750 nm. Wide-field fluorescence images were excited at 640 nm and registered between 660 and 750 nm. High resolution confocal reflectance and fluorescence images were excited at 642 nm. Confocal fluorescence images were acquired between 670 nm and 710 nm. After imaging, the specimens were processed for hematoxylin and eosin (H&E) histopathology. Histological slides were compared with wide-field and high-resolution optical images to evaluate correlation of tumor boundaries and cellular morphology, respectively. Fluorescence polarization imaging identified the location, size, and shape of the tumor in all the cases investigated. Averaged fluorescence polarization values of tumor were higher as compared to normal tissue. Statistical analysis confirmed the significance of these differences. Fluorescence confocal imaging enabled cellular-level resolution. Evaluation and statistical analysis of MB fluorescence polarization values registered from single tumor and normal cells demonstrated higher fluorescence polarization from cancer. Wide-field high-resolution fluorescence and fluorescence polarization imaging shows promise for intraoperative delineation of breast cancers.
Nonmelanoma skin cancers are the most common form of cancer. Continuous wave terahertz imaging has the
potential to differentiate between nonmelanoma skin cancers and normal skin. Terahertz imaging is non-ionizing
and offers a high sensitivity to water content. Contrast between cancerous and normal tissue in transmission mode
has already been demonstrated using a continuous wave terahertz system. The aim of this experiment was to
implement a system that is capable of reflection modality imaging of nonmelanoma skin cancers. Fresh excisions of
skin cancer specimens were obtained from Mohs surgeries for this study. A CO2 optically pumped far-infrared
molecular gas laser was used for illuminating the tissue at 584 GHz. The reflected signal was detected using a liquid
Helium cooled Silicon bolometer. The terahertz images were compared with sample histology. The terahertz
reflection images exhibit some artifacts that can hamper the specificity. The beam waist at the sample plane was
measured to be 0.57 mm, and the system's signal-to-noise ratio was measured to be 65 dB.
Brain tumors cause significant morbidity and mortality even when benign. Completeness of resection of brain tumors improves quality of life and survival; however, that is often difficult to accomplish. The goal of this study was to evaluate the feasibility of using multimodal confocal imaging for intraoperative detection of brain neoplasms. We have imaged different types of benign and malignant, primary and metastatic brain tumors. We correlated optical images with histopathology and evaluated the possibility of interpreting confocal images in a manner similar to pathology. Surgical specimens were briefly stained in 0.05 mg/ml aqueous solution of methylene blue (MB) and imaged using a multimodal confocal microscope. Reflectance and fluorescence signals of MB were excited at 642 nm. Fluorescence emission of MB was registered between 670 and 710 nm. After imaging, tissues were processed for hematoxylin and eosin (H&E) histopathology. The results of comparison demonstrate good correlation between fluorescence images and histopathology. Reflectance images provide information about morphology and vascularity of the specimens, complementary to that provided by fluorescence images. Multimodal confocal imaging has the potential to aid in the intraoperative detection of microscopic deposits of brain neoplasms. The application of this technique may improve completeness of resection and increase patient survival.
Continuous wave terahertz imaging has the potential for diagnosing and delineating skin cancers. While contrast has
been observed between cancerous and normal tissue at terahertz frequencies, the source mechanism behind this contrast
is not clearly understood.1Transmission measurements of 240μm thick sections of nonmelanoma skin cancer were taken
at two frequencies of 1.39 THz and 1.63 THz that lie within and outside the tryptophan absorption band, respectively.
Two CO2 pumped Far-Infrared molecular gas lasers were used for illuminating the tissue while the transmitted signals
were detected using a liquid Helium cooled Silicon bolometer. At both THz frequencies 2-dimensional THz transmission
images of nonmelanoma skin cancers were acquired with better than 0.5mm spatial resolution. The resulting images
were compared to the sample histology and showed a correlation between cancerous tissue and decreased transmission.
The results of the imaging experiments will be presented and discussed.
Skin cancer is the most common form of human cancer. Its early diagnosis and timely treatment is of paramount importance for dermatology and surgical oncology. In this study, we evaluate the use of reflectance and fluorescence confocal microscopy for detecting skin cancers in an in-vivo trial with B16F10 melanoma and SCCVII squamous cell carcinoma in mice. For the experiments, the mice are anesthetized, then the tumors are infiltrated with aqueous solution of methylene blue and imaged. Reflectance images are acquired at 658 nm. Fluorescence is excited at 658 nm and registered in the range between 690 and 710 nm. After imaging, the mice are sacrificed. The tumors are excised and processed for hematoxylin and eosin histopathology, which is compared to the optical images. The results of the study indicate that in-vivo reflectance images provide valuable information on vascularization of the tumor, whereas the fluorescence images mimic the structural features seen in histopathology. Simultaneous dye-enhanced reflectance and fluorescence confocal microscopy shows promise for the detection, demarcation, and noninvasive monitoring of skin cancer development.
Continuous wave terahertz imaging has the potential to offer a safe, non-invasive medical imaging modality for detecting
different types of human cancers. The aim of this study was to identify intrinsic biomarkers for non-melanoma skin
cancer and their absorption frequencies. Knowledge of these frequencies is a prerequisite for the optimal development of
a continuous wave terahertz imaging system for detecting different types of skin cancers. The absorption characteristics
of skin constituents were studied between 20 and 100 cm-1 (0.6 THz - 3 THz). Terahertz radiation is highly absorbed by
water. Thus, the high water content of human tissue necessitates a reflection based imaging modality. To demonstrate a
reflection based, high resolution, terahertz imaging system, a prototype imaging system was constructed at 1.56 THz.
The system resolution was determined to be 0.5 mm and the system signal to noise ratio was found to be 70 dB. Data
from the terahertz spectroscopy experiments and reflection based terahertz images at 1.56 THz are presented.
It has been known for many years that low level laser (or light) therapy (LLLT) can ameliorate the pain, swelling
and inflammation associated with various forms of arthritis. Light is absorbed by mitochondrial chromophores
leading to an increase in ATP, reactive oxygen species and/or cyclic AMP production and consequent gene
transcription via activation of transcription factors. However, despite many reports about the positive effects of
LLLT in medicine, its use remains controversial. Our laboratory has developed animal models designed to
objectively quantify response to LLLT and compare different light delivery regimens. In the arthritis model we
inject zymosan into rat knee joints to induce inflammatory arthritis. We have compared illumination regimens
consisting of a high and low fluence (3 J/cm2 and 30 J/cm2), delivered at a high and low irradiance (5 mW/cm2 and 50 mW/cm2) using 810-nm laser light daily for 5 days, with the effect of conventional corticosteroid
(dexamethasone) therapy. Results indicated that illumination with 810-nm laser is highly effective (almost as good
as dexamethasone) at reducing swelling and that longer illumination time was more important in determining
effectiveness than either total fluence delivered or irradiance. Experiments carried out using 810-nm LLLT on
excisional wound healing in mice also confirmed the importance of longer illumination times. These data will be of
value in designing clinical trials of LLLT.
Nonmelanoma skin cancer is the most common form of human cancer, often resulting in high morbidity. Low visual contrast of these tumors makes their delineation a challenging problem. Employing a linearly polarized monochromatic light source and a wide-field CCD camera, we have developed a technique for fluorescence polarization imaging of the nonmelanoma cancers stained using antibiotics from the tetracycline family. To determine the feasibility of the method, fluorescence polarization images of 86 thick, fresh cancer excisions were studied. We found that the level of endogenous fluorescence polarization was much lower than that of exogenous, and that the average values of fluorescence polarization of tetracycline derivatives were significantly higher in cancerous as compared to normal tissue. Out of 86 tumors [54 stained in demeclocycline (DMN) and 32 in tetracycline (TCN)], in 79 cases (51—DMN, 28—TCN) the location, size, and shape of the lesions were identified accurately. The results of this trial indicate that nonmelanoma skin tumors can be distinguished from healthy tissue based on the differences in exogenous fluorescence polarization of TCN and/or DMN. Therefore, the developed technique can provide an important new tool for image-guided cancer surgery.
Differences in absorption and/or scattering of cancerous and normal skin have the potential to provide a basis for noninvasive cancer detection. In this study, we have determined and compared the in vitro optical properties of human epidermis, dermis, and subcutaneous fat with those of nonmelanoma skin cancers in the spectral range from 370 to 1600 nm. Fresh specimens of normal and cancerous human skin were obtained from surgeries. The samples were rinsed in saline solution and sectioned. Diffuse reflectance and total transmittance were measured using an integrating sphere spectrophotometer. Absorption and reduced scattering coefficients were calculated from the measured quantities using an inverse Monte Carlo technique. The differences between optical properties of each normal tissue-cancer pair were statistically analyzed. The results indicate that there are significant differences in the scattering of cancerous and healthy tissues in the spectral range from 1050 to 1400 nm. In this spectral region, the scattering of cancerous lesions is consistently lower than that of normal tissues, whereas absorption does not differ significantly, with the exception of nodular basal cell carcinomas (BCC). Nodular BCCs exhibit significantly lower absorption as compared to normal skin. Therefore, the spectral range between 1050 and 1400 nm appears to be optimal for nonmelanoma skin cancer detection.
It has been known for many years that low levels of laser or non-coherent light (LLLT) accelerate some phases of wound healing. LLLT can stimulate fibroblast and keratinocyte proliferation and migration. It is thought to work via light absorption by mitochondrial chromophores leading to an increase in ATP, reactive oxygen species and consequent gene transcription. However, despite many reports about the positive effects of LLLT on wound healing, its use remains controversial. Our laboratory has developed a model of a full thickness excisional wound in mice that allows quantitative and reproducible light dose healing response curves to be generated. We have found a biphasic dose response curve with a maximum positive effect at 2 J/cm2 of 635-nm light and successively lower beneficial effects from 3-25 J/cm2, the effect is diminished at doses below 2J/cm2 and gradually reaches control healing levels. At light doses above 25 J/cm2 healing is actually worse than controls. The two most effective wavelengths of light were found to be 635 and 820-nm. We found no difference between filtered 635±15-nm light from a lamp and 633-nm light from a HeNe laser. The strain and age of the mouse affected the magnitude of the effect. Light treated wounds start to contract after illumination while control wounds initially expand for the first 24 hours. Our hypothesis is that a single brief light exposure soon after wounding affects fibroblast cells in the margins of the wound. Cells may be induced to proliferate, migrate and assume a myofibroblast phenotype. Our future work will be focused on understanding the mechanisms underlying effects of light on wound healing processes.
Early detection and precise excision of neoplasms are imperative requirements for successful cancer treatment. In this study we evaluated the use of dye-enhanced confocal microscopy as an optical pathology tool in the ex vivo trial with fresh thick non-melanoma
skin cancer excisions and in vivo trial with B16F10 melanoma cancer in mice. For the experiments the tumors were rapidly stained using aqueous solutions of either toluidine blue or methylene blue and imaged using multimodal confocal microscope. Reflectance images
were acquired at the wavelengths of 630nm and 650 nm. Fluorescence was excited at 630 nm and 650 nm. Fluorescence emission was registered in the range between 680 nm and 710 nm. The images were compared to the corresponding en face frozen H&E sections. The
results of the study indicate confocal images of stained cancerous tissue closely resemble corresponding H&E sections both in vivo and in vitro. This remarkable similarity enables interpretation of confocal images in a manner similar to that of histopathology. The
developed technique may provide an efficient real-time optical tool for detecting skin pathology.
Multispectral polarized light imaging (MSPLI) enables rapid inspection of a superficial tissue layer over large surfaces, but does not provide information on cellular microstructure. Confocal microscopy (CM) allows imaging within turbid media with resolution comparable to that of histology, but suffers from a small field of view. In practice, pathologists use microscopes at low and high power to view tumor margins and cell features, respectively. Therefore, we study the combination of CM and MSPLI for demarcation of nonmelanoma skin cancers. Freshly excised thick skin samples with nonmelanoma cancers are rapidly stained with either toluidine or methylene blue dyes, rinsed in acetic acid, and imaged using MSPLI and CM. MSPLI is performed at 630, 660, and 750 nm. The same specimens are imaged by reflectance CM at 630, 660, and 830 nm. Results indicate that CM and MSPLI images are in good correlation with histopathology. Cytological features are identified by CM, and tumor margins are delineated by MSPLI. A combination of MSPLI and CM appears to be complementary. This combined in situ technique has potential to guide cancer surgery more rapidly and at lower cost than conventional histopathology.
More than two million cases of nonmelanoma skin cancers are diagnosed every year. Therefore, there is a strong need for practical, reliable, rapid, and precise methods for tumor delineation, to guide surgery and other treatments of skin cancer. Once developed, such methods may be useful for squamous cell carcinomas of other organs. Non-invasive optical imaging techniques including polarization sensitive reflectance and fluorescence imaging were evaluated for the demarcation of nonmelanoma skin tumors. Thick freshly excised tumor specimens obtained from Mohs surgery were used for the experiments. Imaging was performed using linearly polarized incident light in the visible and near infrared spectral range from 577 nm to 750 nm. Non-toxic absorbing and fluorescent dyes (Toluidine Blue O, Methylene Blue) were employed to enhance tumor contrast in the images. The images were acquired using the remitted light polarized in the directions parallel and perpendicular to the polarization of incident light. Reflectance and fluorescence polarization images were evaluated. The data were processed and analyzed for dependence of the remitted light polarization on the tissue type (cancerous/normal). The data obtained so far from fresh tumor specimens in vitro using dye-enhanced polarized light reflectance, and exogenous fluorescence polarization imaging suggest that optical mapping can become a valuable guidance tool in nonmelanoma cancer surgery.
Many diseased states of the brain can result in the displacement of brain tissues and restrict cerebral blood flow, disrupting function in a life-threatening manner. Clinical examples where displacements are observed include venous thromboses, hematomas, strokes, tumors, abscesses, and, particularly, brain edema. For the latter, the brain tissue swells, displacing the cerebral spinal fluid (CSF) layer that surrounds it, eventually pressing itself against the skull. Under such conditions, catheters are often inserted into the brain's ventricles or the subarachnoid space to monitor increased pressure. These are invasive procedures that incur increased risk of infection and consequently are used reluctantly by clinicians. Recent studies in the field of biomedical optics have suggested that the presence or absence of the CSF layer can lead to dramatic changes in NIR signals obtained from diffuse reflectance measurements around the head. In this study, we consider how this sensitivity of NIR signals to CSF might be exploited to non-invasively monitor the onset and resolution of brain edema.
A novel approach to the quantitative image reconstruction in diffuse optical tomography is proposed. The special structure of the transport equation is used to formulate the iterative image reconstruction algorithm as a process updating the estimates of the optical properties from the solution of an intermediate tomographic problem The ability of the technique to reconstruct simultaneously maps of both absorption and reduced scattering coefficients in 2D geometry is demonstrated using simulated frequency-domain data. The potential advantages of the new approach include its ability to fully retain the non-linear character of the inverse problem while at the same time avoiding either gradient or Jacobian calculations and eliminating the need in an additional regularization mechanism.
Proc. SPIE. 3597, Optical Tomography and Spectroscopy of Tissue III
KEYWORDS: Data modeling, Modulation, Optical properties, Scattering, Sensors, Diffusion, Phase shift keying, Monte Carlo methods, Tissue optics, Signal detection
Optical reflectance spectroscopy of turbid media is one of promising novel techniques for diagnostics and monitoring of biological tissues. A widely used approach in the realization of this technique is the application of photon-density waves. Until recently, the frequency-domain techniques have been limited to probing relatively large volumes of tissue. Main reason for this limitation was almost exclusive use of the diffusion approximation for the interpretation of the measured data. The diffusion model becomes invalid for source-detector separations less than several transport mean free paths (approximately 5-7 mm for most of the soft tissues in the near-infrared spectral range). We have developed an inverse technique that allows data processing even at small source-detector separations. This technique is based on a frequency-domain-optiinized Monte Carlo algorithm as a light propagation model. A specially designed acceleration scheme affords using the technique as a forward method in an iterative inverse algorithm. In this paper, a description of the technique is given, and results of simulations and preliminary phantom experiments are presented.
We determined and compared to the optical properties of five samples obtained from xenotransplanted chick embryos chorio- allantoic membrane tumors, five samples obtained from xenotransplanted chick embryos yolk sac membrane tumors, and five samples of concentrated tumor cells suspension (small cell lung carcinoma OAT 75). The absorption coefficient (mu) a, the scattering coefficient (mu) s, the anisotropy factor g, and the reduced scattering coefficient (mu) s' were evaluated in the spectral range from 600 nm to 900 nm with a step width of 10 nm from double integrating sphere measurements using an inverse Monte Carlo technique. The results have shown that the optical properties of the concentrated tumor cell suspension are similar to those of the chorio-allantoic membrane tumor, but are essentially different from the optical properties of the yolk sac membrane tumor. Cell vitality tests have shown that the cells were alive during and after the experiments. Therefore, the tumor cell suspension can serve as an optical model of the chorio-allantoic membrane tumor in situ for testing and developing novel diagnostic and therapeutic techniques.
The optical properties of seven native, seven laser-coagulated (Nd:YAG (lambda) equals 1064 nm, 11 W, 15 min, T less than or equal to 90 degrees Celsius), and seven thermally coagulated (60 min, T less than or equal to 80 degrees Celsius) samples of bovine myocardium were determined. The absorption coefficient (mu) a, the scattering coefficient (mu) s, and the anisotropy factor g were obtained in the spectral range from 1000 nm to 1500 nm from double integrating sphere measurements using an inverse Monte Carlo technique. The results indicate that both, laser coagulation and thermal coagulation increase the (mu) a and (mu) s values by a factor of 2 and 4, respectively, while g is not significantly changed. Conclusion: Thermal denaturation leads to significant changes in the optical properties of bovine myocardium. The changes are comparable to those induced by laser coagulation.
In optics of biotissues it is often required to determine the radiance distribution function of a finite beam propagating through a turbid medium. One particular instance of this problem is the determination of the signal registered by a detector with a limited field of view, placed on the axis of beam propagation. Since most biological tissues exhibit a high anisotropy factor of scattering, the problem can be approached by applying the small-angle approximation of the radiative transfer theory. We have developed a technique for solving the small-angle problem in case of the Henyey-Greenstein phase function. The technique accounts for an arbitrary spatial and angular profile of the incident beam. The results of the numerical tests have shown a good agreement with the predictions of forward Monte Carlo simulations for a variety of optical properties typical for biological tissues.
The development of time-resolved optical diagnostic techniques for biomedical applications requires an accurate description of the time-dependent photon propagation in tissues. In many applications the diffusion approximation is used for this purpose. However, in case of a highly anisotropic scattering and in the vicinity of light sources the diffusion equation becomes inadequate. To overcome this limitation, we introduce another approximation of the time- dependent radiative transfer equation. The approximation is based on the assumption that the scattering phase function of the medium is strongly forward-peaked, which has been established for a variety of tissues. We show that in this case, the integro-differential time-dependent transfer equation can be reduced to a partial differential equation. Furthermore, we demonstrate that this approximate equation is valid at much shorter distances from the source than the diffusion equation. At the same time, this approach is amenable for a combination with an inverse technique in order to determine the optical properties of the medium from a time- or frequency-resolved experiment.
The laser-induced interstitial thermo-therapy of brain tumors requires an exact therapy planning. Therefore, the knowledge of the optical properties of native (na) and coagulated (co) tissue structures is important. In this study the optical properties of native and thermally coagulated (2 h, 80 degree Celsius) human white (n equals 14; na equals 7, co equals 7) and gray matter (n equals 14; na equals 7, co equals 7) were investigated (spectral range equals 360 - 1100 nm, spectral resolution 20 nm) in vitro using the integrating sphere technique combined with the inverse Monte-Carlo method. The (mu) a of the native gray matter decreased from 0.333 plus or minus 0.219/mm (360 nm) to 0.054 plus or minus 0.069/mm (1100 nm). The (mu) s varied between 14.13 plus or minus 4.26/mm (360 nm) and 5.53 plus or minus 1.80/mm (1100 nm). The g-value increased from 0.818 plus or minus 0.093 (360 nm) to 0.904 plus or minus 0.051 (1100 nm). Coagulation increased (mu) a and (mu) s by a factor up to 3 and g up to 16%. White matter exhibited a (mu) a between 0.253 plus or minus 0.055/mm (360 nm) and 0.100 plus or minus 0.052/mm (1100 nm) and a (mu) s between 40.20 plus or minus 9.18/mm (360 nm) and 28.65 plus or minus 6.83/mm (1100 nm). The g-value varied between 0.702 plus or minus 0.093 (360 nm) and 0.886 plus or minus 0.012 (1100 nm). Coagulation increased (mu) a up to a factor of three and the g-value up to 14% while the increase (approximately 1.25 fold) of (mu) s was not significant. We conclude the optical properties of human brain tissue change significantly due to thermal denaturation.
We investigated the impact of the scattering phase function approximation on the optical properties of whole human blood determined from integrating sphere measurements using an inverse Monte Carlo technique. The diffuse reflectance Rd and the total transmittance Tt ((lambda) equals 633nm) of the whole blood samples were measured with a double integrating sphere equipment. The experimental scattering phase functions of the highly diluted blood samples were measured with a goniophotometer. We approximated the experimental scattering phase function with Mie, Gegenbauer kernel (GKPF), and Henyey-Green (HGPF) phase functions to pre-set the anisotropy factor (mu) for the inverse problem. We have employed HGPF, GKPF, and MPF approximations in the inverse Monte Carlo procedure to derive the absorption coefficient (mu) a and the scattering coefficient (mu) s. The results show significant difference in the final estimates of (mu) s. 12
We determine the optical properties of whole blood samples in the near infrared spectral range from double integrating sphere measurements using an inverse Monte Carlo technique. The measured values included the diffuse reflectance, the total transmittance, and the collimated transmittance. From these data, the absorption coefficient, the scattering coefficient, and the anisotropy factor were derived. The spectral range investigated extended from 700 nm to 1200 nm. It was found that the optical properties of blood were substantially different from the respective data for other relevant human tissues known so far. In addition, we analyzed the effect of the scattering phase function approximation on the resulting estimates of the optical properties. The Henyev-Greenstein and the Gegenbauer kernel phase functions were considered. The calculated angular distributions of scattered light were compared with goniophotometric measurements performed at the wavelength of 633 nm. The data presented in this study prove that the variations of the employed scattering phase function approximation can cause large discrepancies in the derived optical properties. This leads to the conclusion that the exact knowledge of the scattering phase function is required for the precise determination of the optical constants from the double integrating sphere measurements.
The accurate prediction of the temperature and damage distribution, respectively, is a prerequisite for the successful application of the laser-induced interstitial thermotherapy of brain tumors. For the precise calculation of the resulting damage, the exact structure of the tissue in the region of interest has to be taken into account. We used the MR images of the respective brain tumor as the starting point for the computation of the light, the heat, and the consecutive damage distribution within the individual tissue structures selected for treatment. The simulation technique is based on the combination of the Monte Carlo method to calculate the light distribution and a finite difference alternative directions implicit scheme to solve the heat transfer equation. The model accounts for the specific geometry of the applicator, the changes of the optical and thermal tissue properties during the heating process, and the blood perfusion. The boundary conditions are determined by the thermal exchange between the tissue, the applicator, and the ambient medium. The predictions of the model were compared with the MR image of the resulting laser lesion induced in a patient. The relative error in the determination of the size of the damage area was found to be within 10 per cent in the direction of the applicator axis and within 20 per cent in the radial direction.
The time-dependent equation of the radiative transport is reduced to the stationary one for the case of a radiation source being modulated by a harmonic frequency. A Monte Carlo scheme is suggested to solve the resulting equation. The technique avoids tracking the time-histories of each individual photon and allows us to take the finite single-scattering transient time into account. The algorithm directly estimates the quantities being relevant to frequency-domain measurements. A single set of photon trajectories can be used to compute the modulation and the phase of the scattered radiation at different modulation frequencies. The results of the Monte Carlo simulations are compared to predictions of the rigorous radiative transport theory and the diffusion approximation. It is found that the Monte Carlo technique provides a good agreement with the transport theory whereas the accuracy of the diffusion approximation decreases with growth of the modulation frequency. In addition, the technique is used to study the effect of the finite single-scattering transient time on the resulting modulation and phase distribution of the diffusely reflected radiation. It is shown that even a transient time as short at 0.1 ps can significantly affect the reflected signal from a medium presenting optical properties similar to those of biological tissues in the near-infrared spectral range.
In this work, we present a comparison of two approaches (the rigorous transfer equation and its diffusion approximation) to describe the frequency dependence of the modulation and the phase shift of scattered light for the problem of diffuse reflection form a semi-infinite medium with isotropic scattering. It is shown that both approaches lead to the same results, when the modulation frequency of the incident light is low with respect to the inverse time-of-flight between two interaction sites. At increased frequencies, however, these two models reveal differences in their predictions. Besides, the diffusion approximation fails even qualitatively to describe angular dependence of the modulation and the phase shift of back scattered radiation. The photon migration process in tissue is also influenced by the finite (non-zero) life-time of photons in the virtually absorbed state during the scattering process. Most of the existing models assume that this life-time is neglectably short. However, in dense scattering media, like in most biological tissues, this time can be comparable with the time-of-flight between two interaction sites. We also investigated this effect on the frequency dependence of the modulation and the phase shift. The results let us conclude that both -- the diffusion approximation and the assumption of short life-times in the virtually absorbed state -- should be applied with caution when using frequency domain data to determine optical properties of biological tissues especially when using high modulation frequencies. This is also true for the application of this technique in optical tomography.
There is a defmite need to determine accurately optical parameters of tissues. It is required either when planning a laser treatment or trying to make diagnostic conclusions, based on differences in optical properties of healthy and pathological tissues. Scattering phase function is an important characteristic of medium optical properties1.Knowledge of scattering phase function of epidermal layers is necessary for direct calculations of light propagation through skin2. Changes in angular properties of scattered light may reflect pathology of tissue and can be used in investigation of skin diseases, like psoriasis3. Besides the self4mportance measurements of scattering phase function can be used in combination with integrating sphere measurements to determine absorption and scattering coefficients of tissue4. This combination can also be useful for testing of photoprotectors. Direct gomometric measurements of the angular distribution of radiation, scattered by human samples have been reported by several authors5'6. However, known literature data on angular scattering properties of human epidermis are still liniited7'8. In present paper we concentrate on outermost epidermal layers and report measurements, performed on samples, obtained by the novel technique3. We investigated also influence of various chemical's applications to skin surface on scattering phase function of epidermal layers.
Optical models of crystalline lens were analyzed, to increase understanding of high transparency and loss of it with aging and for pathology tissue. Experimental and calculated human crystalline lens spectra in a wide range of wavelengths are presented. Calculations were made for the model of short-term- order scatterers, taking into account interference effects of scatterers and specific features of absorbing species, such as protein-bound tryptophan, kynurenine, and age-related chromophores leading to generalized yellowing of the lens proteins. Satisfactory qualitative coincidence of calculated spectra with spectrophotometric data for isolated human lens is shown. Measured angular dependencies of scattering matrix elements are sensitive to modification related to aging and cataract appearance.
Elastic light scattering and Raman light scattering applied to the same eye-lens have been used, respectively, to extract information on the spatial variations in the intensity of scattered light and the protein content. A combination of the results allows one to obtain the distribution of the scattering coefficient, the size of the scattering particles and the molecular weight of the scattering particles. The design of the light scattering set up is such that the results can be compared with those obtained with a `Scheimpflug' camera. The increase in light scattering at the anterior and posterior cortex in young (< 20 - 30 yrs.) eye-lenses is in accordance with the theory for short-range crystalline order in eye-lenses.
New technology for the definition of the skin epidermis optical parameters, which includes upper layers of epidermis stripping, diffuse reflection, transmission, and angular measurement of scattered light by stripping samples, calculations using 4-flux Kubelka-Munk approximation, have been obtained. This technology was successfully used for depth dependence monitoring of epidermis optical parameters, and for laser light dosimetry in percutaneous irradiation of blood and laser PUVA therapy.
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