We have applied a CNN to preprocess Raman spectra from fresh tissue samples from brain tumors. The neural network can handle the variations that occur naturally, which enables explorative data analysis methods such as PCA.
Laser Doppler flowmetry (LDF) has been adapted for long-term neurointensive care monitoring of cerebral microcirculation at two sites. The invasive probe was successfully used in a 10-day recording in one patient with subarachnoid hemorrhage (SAH).
In the routine of stereotactic biopsy on suspected tumors located deep in the brain or patients with multiple lesions, tissue samples are harvested to determine the type of malignancy. Biopsies are taken from pre-calculated positions based on the preoperative radiologic images susceptible to brain shift. In such cases the biopsy procedure may need to be repeated leading to a longer operation time. To provide guidance for targeting diagnostic tumor tissue and to avoid vessel rupture on the insertion path of the tumor, an application specific fiber optic probe was developed. The setup incorporated spectroscopy for 5-aminolevulinic acid induced protopophyrin IX (PpIX) fluorescence in the tumor and laser Doppler for measuring microvascular blood flow which recorded backscattered light (TLI) at 780 nm and blood perfusion. The recorded signals were compared to the histopathologic diagnosis of the tissue samples (n=16) and to the preoperative radiologic images. All together 146 fluorescence and 276 laser Doppler signals were recorded along 5 trajectories in 4 patients. On all occasions strong PpIX fluorescence peaks were visible during real-time guidance. Comparing the gliotic tumor marginal zone with the tumor, the PpIX (51 vs. 528 a.u., [0-1790], p < 0.05) was higher and TLI (2.9 vs. 2.0 a.u., [0-4.1], p < 0.05) was lower in tumor. The autofluorescence (104 vs.70 a.u., [0-442], p > 0.05) and blood perfusion (8.3 vs. 17 a.u., [0-254], p > 0.05) were not significantly different. In conclusion, the optical guidance probe made real-time tumor detection and vessel tracking possible during the stereotactic biopsy procedures. Moreover, the fluorescence and blood perfusion in the tumor could be studied at controlled positions in the brain and the tumor.
Image processing algorithms in pathology commonly include automated decision points such as classifications. While
this enables efficient automation, there is also a risk that errors are induced. A different paradigm is to use image
processing for enhancements without introducing explicit classifications. Such enhancements can help pathologists to
increase efficiency without sacrificing accuracy. In our work, this paradigm has been applied to Ki-67 hot spot detection.
Ki-67 scoring is a routine analysis to quantify the proliferation rate of tumor cells. Cell counting in the hot spot, the
region of highest concentration of positive tumor cells, is a method increasingly used in clinical routine. An obstacle for
this method is that while hot spot selection is a task suitable for low magnification, high magnification is needed to
discern positive nuclei, thus the pathologist must perform many zooming operations. We propose to address this issue by
an image processing method that increases the visibility of the positive nuclei at low magnification levels. This tool
displays the modified version at low magnification, while gradually blending into the original image at high
magnification. The tool was evaluated in a feasibility study with four pathologists targeting routine clinical use. In a task
to compare hot spot concentrations, the average accuracy was 75±4.1% using the tool and 69±4.6% without it (n=4). Feedback on the system, gathered from an observer study, indicate that the pathologists found the tool useful and fitting in their existing diagnostic process. The pathologists judged the tool to be feasible for implementation in clinical routine.
Fluorescence guidance using 5-aminolevulinic acid (5-ALA) for brain tumor resection is a recent technique applied to the highly malignant brain tumors. Five-ALA accumulates as protoporphyrin IX fluorophore in the tumor cells in different concentrations depending on the tumor environment and cell properties. Our group has developed a fluorescence spectroscopy system used with a hand-held probe intra-operatively. The system has shown improvement of fluorescence detection and allows quantification that preliminarily correlates with tumor malignancy grade during surgery. However, quantification of fluorescence is affected by several factors including the initial fluorophore concentration, photobleaching due to operating lamps and attenuation from the blood. Accordingly, an optical phantom was developed to enable controlled fluorescence measurements and evaluation of the system outside of the surgical procedure. The phantom mimicked the optical properties of glioma at the specific fluorescence excitation wavelength when different concentrations of the fluorophore were included in the phantom. To allow evaluation of photobleaching, kinetics of fluorophore molecules in the phantom was restricted by solidifying the phantoms. Moreover, a model for tissue autofluorescence was added. The fluorescence intensity’s correlation with fluorophore concentration in addition to the photobleaching properties were investigated in the phantoms and were compared to the clinical data measured on the brain tumor.
The principles of cancer treatment has for years been radical resection of the primary tumor. In the oncologic surgeries where the affected cancer site is close to the lymphatic system, it is as important to detect the draining lymph nodes for metastasis (lymph node mapping). As a replacement for conventional radioactive labeling, indocyanine green (ICG) has shown successful results in lymph node mapping; however, most of the ICG fluorescence detection techniques developed are based on camera imaging. In this work, fluorescence spectroscopy using a fiber-optical probe was evaluated on a tissue-like ICG phantom with ICG concentrations of 6-64 μM and on breast tissue from five patients. Fiber-optical based spectroscopy was able to detect ICG fluorescence at low intensities; therefore, it is expected to increase the detection threshold of the conventional imaging systems when used intraoperatively. The probe allows spectral characterization of the fluorescence and navigation in the tissue as opposed to camera imaging which is limited to the view on the surface of the tissue.
Fluorescence guidance in brain tumor resection is performed intra-operatively where bleeding is included. When using
fiber-optical probes, the transmission of light to and from the tissue is totally or partially blocked if a small amount of
blood appears in front of the probe. Sometimes even after rinsing with saline, the remnant blood cells on the optical
probe head, disturb the measurements. In such a case, the corresponding spectrum cannot be reliably quantified and is
therefore discarded. The optimal case would be to calculate and take out the blood effect systematically from the
collected signals. However, the first step is to study the pattern of blood interference in the fluorescence spectrum. In this
study, a fiber-optical based fluorescence spectroscopy system with a laser excitation light of 405 nm (1.4 J/cm2) was
used during fluorescence guided brain tumor resection using 5-aminolevulinic acid (5-ALA). The blood interference
pattern in the fluorescence spectrum collected from the brain was studied in two patients. The operation situation was
modeled in the laboratory by placing blood drops from the finger tip on the skin of forearm and the data was compared to
the brain in vivo measurements. Additionally, a theoretical model was developed to simulate the blood interference
pattern on the skin autofluorescence. The blood affects the collected fluorescence intensity and leaves traces of oxy and
deoxy-hemoglobin absorption peaks. According to the developed theoretical model, the autofluorescence signal is
considered to be totally blocked by an approximately 500 μm thick blood layer.
An electrode with adjacent optical fibers for measurements during navigation and radio frequency lesioning in the brain is modeled for Monte Carlo simulations of light transport in brain tissue. Relative reflected light intensity at 780 nm, I780, from this electrode and probes with identical fiber configuration are simulated using the intensity from native white matter as reference. Models are made of homogeneous native and coagulated gray, thalamus, and white matter as well as blood. Dual layer models, including models with a layer of cerebrospinal fluid between the fibers and the brain tissue, are also made. Simulated I780 was 0.16 for gray matter, 0.67 for coagulate gray matter, 0.36 for thalamus, 0.39 for coagulated thalamus, unity for white matter, 0.70 for coagulated white matter, and 0.24 for blood. Thalamic matter is also found to reflect more light than gray matter and less than white matter in clinical studies. In conclusion, the reflected light intensity can be used to differentiate between gray and white matter during navigation. Furthermore, coagulation of light gray tissue, such as the thalamus, might be difficult to detect using I780, but coagulation in darker gray tissue should result in a rapid increase of I780.
The highly malignant brain tumor, glioblastoma multiforme (GBM), is difficult to fully delineate during surgical
resection due to its infiltrative ingrowth and morphological similarities to surrounding functioning brain tissue.
Selectiveness of GBM to 5-aminolevulinic acid (5-ALA) induced protoporphyrin IX (PpIX) is reported by other
researchers to visualize tumor margins under blue light microscopy. To allow objective detection of GBM, a compact
and portable fiber optic based fluorescence spectroscopy system is developed. This system is able to deliver excitation
laser light (405 nm) in both the continuous and pulsed mode. PpIX fluorescence peaks are detected at 635 and 704 nm,
using a fiber-coupled spectrometer. It is necessary to optimize the detection efficiency of the system as the PpIX quickly
photobleaches during the laser illumination. A light dose of 2.5 mJ (fluence rate = 9 mJ/mm2) is experimentally approved
to excite an acceptable level of fluourescence signal arising from glioblastoma. In pulsed illumination mode, an
excitation dose of 2.5 mJ, with a dark interval of 0.5 s (duty cycle 50%) shows a significantly shorter photobleaching
time in comparison to the continuous illumination mode with the same laser power (p < 0.05). To avoid photobleaching
(the remaining signal is more than 90% of its initial value) when measuring with 2.5 mJ delivered energy, the time for
continuous and pulsed illumination should be restricted to 2.5 and 1.1 s, respectively.
The highly malignant brain tumor, glioblastoma multiforme, is difficult to totally resect without aid due to its infiltrative
way of growing and its morphological similarities to surrounding functioning brain under direct vision in the operating
field. The need for an inexpensive and robust real-time visualizing system for resection guiding in neurosurgery has been
formulated by research groups all over the world. The main goal is to develop a system that helps the neurosurgeon to
make decisions during the surgical procedure.
A compact fiber optic system using fluorescence spectroscopy has been developed for guiding neurosurgical resections.
The system is based on a high power light emitting diode at 395 nm and a spectrometer. A fiber bundle arrangement is
used to guide the excitation light and fluorescence light between the instrument and the tissue target. The system is
controlled through a computer interface and software package especially developed for the application. This robust and
simple instrument has been evaluated in vivo both on healthy skin but also during a neurosurgical resection procedure.
Before surgery the patient received orally a low dose of 5-aminolevulinic acid, converted to the fluorescence tumor
marker protoporphyrin IX in the malignant cells. Preliminary results indicate that PpIX fluorescence and brain tissue
autofluorescence can be recorded with the help of the developed system intraoperatively during resection of glioblastoma
multiforme.
Laser Doppler perfusion imaging (LDPI) enables superficial tissue perfusion assessment, but is sensitive to tissue motion not related to blood cells. The aim was to investigate if a polarization technique could reduce movement-induced artifacts. A linearly polarized laser and a cross-polarized filter, placed in front of the detectors, were used to block specular reflection. Measurements were performed with, and without, the polarization filter, at a single site during horizontal and vertical movement of skin tissue (index finger, twelve subjects, n=112) and of a flow model (n=432), with varying surface structures. Measurements were repeated during different flow conditions and at increased skin specular reflection. Statistical analysis was performed using ANOVA models. The perfusion signal was lower (p<0.001, skin and p<0.05, flow model) using the polarization filter, due to movement artifact reduction. No significant influence from surface structure was found when using the polarization filter. Movement artifacts were lower (p<0.05) in the vertical movement direction, however, depending on flow conditions for skin measurements. Increased skin specular reflection gave rise to large movement artifacts without the polarization filter. In conclusion, the polarized LDPI technique reduces movement artifacts and is particularly appropriate when assessing, e.g., ulcers and burns, where specular reflection is high.
In numerous medical and scientific fields, knowledge of the optical properties of tissues can be applied. Among many different ways of determining the optical properties of turbid media; integrating sphere measurements are widely used. However, this technique is associated with bulky equipment, complicated measuring techniques, interference compensation techniques, and inconvenient sample handling. This paper describes measurements of the optical properties of porcine brain tissue using novel instrumentation for simultaneous absorption and scattering characterization of small turbid samples. The system used measures both angularly and spatially resolved transmission and reflection and is called Combined Angular and Spatially-resolved Head (CASH) sensor. The results compare very well with data obtained with an integrating sphere for well-defined samples. The instrument was shown to be accurate to within 12% for μa, and 1% for μs' in measurements of intralipid-ink samples. The corresponding variations of data were 17%, and 2%, respectively. The reduced scattering coefficient for porcine white matter was measured to be 100 cm-1, while the value for coagulated brain tissue was 65 cm-1. The corresponding absorption coefficients were 2 and 3 cm-1, respectively.
Radiofrequency (RF) lesioning in the human brain is one possible surgical therapy for severe pain as well as movement disorders. One obstacle for a safer lesioning procedure is the lack of size monitoring. The aim of this study was to investigate if changes in laser Doppler or intensity signals could be used as markers for size estimation during experimental RF lesioning. A 2 mm in diameter monopolar RF electrode was equipped with optical fibers and connected to a digital laser Doppler system. The optical RF electrode's performance was equal to a standard RF electrode with the same dimensions. An albumin solution with scatterers was used to evaluate the intensity and laser Doppler signal changes during lesioning at 70, 80, and 90 °C. Significant signal changes were found for these three different clot sizes, represented by the temperatures (p<0.05, n=10). The volume, width, and length of the created coagulations were correlated to the intensity signal changes (r=0.88, n=30, p<0.0001) and to the perfusion signal changes (r=0.81, n=30, p<0.0001). Both static and Doppler-shifted light can be used to follow the lesioning procedure as well as being used for lesion size estimation during experimental RF lesioning.
Radio frequency (RF) lesioning in the human brain is a common surgical therapy for relieving severe pain as well as for movement disorders such as Parkinsonia. During the procedure a small electrode is introduced by stereotactic means towards a target area localized by CT or MRI. An RF-current is applied through the electrode tip when positioned in the target area. The tissue in the proximity of the tip is heated by the current and finally coagulated.
The overall aim of this study was to improve the RF-technique and its ability to estimate lesion size by means of optical methods. Therefore, the optical differences between white and gray matter, as well as lesioned and unlesioned tissue were investigated. Reflection spectroscopy measurements in the range of 450-800 nm were conducted on fully anesthetized pigs during stereotactic RF-lesioning (n=6). Light from a tungsten lamp was guided to the electrode tip through optical fibers, inserted along a 2 mm in diameter monopolar RF-electrode. Measurements were performed in steps of 0-10 mm from the target in each hemisphere towards the entry point of the skull. In the central gray of the porcine brain measurements were performed both before and after the creation of a lesion. A total of 55 spectra were collected during this study. Correlation to tissue type was done using post-operative MR-images. The spectral signature for white and gray matter differs significantly for the entire spectral range of 450-800 nm. Pre- and post-lesioning reflection spectroscopy showed the largest differences below 600 and above 620 nm, which implies that lasers within this wavelength range may be useful for in-vivo measurements of tissue optical changes during RF-lesioning.
The microvascular perfusion can be measured using laser Doppler blood flowmetry (LDF), a technique sensitive to the concentration of moving blood cells and their velocity. However, movements of the tissue itself can cause artifacts in the perfusion readings. In a clinical situation, these movement induced artifacts may arise from patient movements or from movements of internal organs e.g. the intestines or the beating heart. Therefore, we have studied how a well-controlled tissue movement affects the LDF signals during different flow conditions and for different surface structures. Tissue perfusion was recorded non-touch in one point using a laser Doppler perfusion imager. During the measurements the object was placed on a shaker that generated the movement (both horizontal and vertical). Measurements were carried out both on DELRIN (polyacetal plastic) and the fingertip, for a wide range of velocities (0-3 cm/s). The influence of the microvascular perfusion was evaluated by occluding the brachial artery as well as blood emptying the finger and by using a flow model. The LDF signals were correlated to the movement. In vivo measurements showed that velocities above 0.8 cm/s gave a significant contribution to the perfusion signal. Corresponding velocities for the DELRIN piece were higher (1.4 - 2.6 cm/s), and dependent on the surface structures and reflecting properties. By reducing the amount of specular reflection the movement influence was substantially lowered.
Stereotactic radiofrequency (RF)-lesioning in the central part of the brain is performed on patients that, for instance, have severe movement or psychiatric disorders. The size of the generated lesion can to some extent be controlled by RF-generator settings such as temperature and time as well as the electrode configuration. Today, MR- imaging and CT are the essential diagnostic methods to confirm the lesion size in vivo. The aim of this study was to investigate whether it is possible to use changes in the reflected light intensity and laser Doppler flowmetry as a marker for size estimation during RF-lesioning.
Laser Doppler perfusion imaging (LDPI) has successfully been used to map the myocardial perfusion on patients undergoing coronary bypass surgery on the arrested heart. The need for intra-operative evaluation of graft function is obvious in routine surgery but even more imperative when adapting new surgical techniques where the procedure is performed on the beating heart. When using LDPI on the beating heart, artifacts originating from the movement of the heart are superimposed on the Doppler signal. We have investigated a method to reduce these artifacts by controlling the sampling sequence with ECG-triggering. The method has been assessed in an animal model on the beating calf heart. After sternotomy, an area covering 1 cm2 was imaged at the anterior wall of the left ventricle. In this area, six perfusion images were captured each of them recorded at fixed, but different time intervals in the cardiac cycle. In addition continuous measurements at one spot was done during 1 - 2 minutes. The signal recorded during pumping action was high compared to measurements performed in the same muscle area during infusion of blood with a syringe pump. Repeated measurements captured at a fixed delay time from the R-peak in the same areas at the same heart frequency showed reproducibility. ECG-triggering of the laser Doppler signal is the first step in our attempts to adapt LDPI to enabling assessment of myocardial perfusion on the beating heart. Further technical achievements and in-vivo investigations are, however, needed and will be performed by our research team in future studies.
Laser Doppler perfusion imaging has been used to assess the myocardium perfusion on the arrested heart during bypass surgery. Twenty-two patients undergoing coronary artery bypass grafting, including usage of the left internal thoracic artery, were included in the study. The anticipated perfusion increase following declamping of the internal thoracic artery was investigated by mapping areas at the size of 10 cm X 11 cm, (n equals 11) and 7 cm X 5 cm (n equals 11). The larger images allowed quantification of blood flow in different regions of the myocardium. The size of the affected area was 32.2 +/- 12.9 cm2 with a total increase of 3.17 +/- 0.75 a.u. (range 0 - 10 a.u.). Corresponding values for areas surrounding the vessels and areas defined as the larger vessels in the myocardium were 29.0 +/- 10.9 cm2 (2.85 +/- 0.57 a.u.) and 3.5 +/- 2.8 cm2 (6.78 +/- 0.18 a.u.). All subjects but two showed a substantial blood flow increase (> 2 a.u.) after release of the clamp. Six subjects had a total increase of at least 4 a.u. Correlation analysis between areas including various number of sites showed an r equals 0.91 (p < 0,0001) or better. In conclusion, laser Doppler perfusion imaging can easily be used intraoperatively in conjunction with bypass surgery. It enables immediate assessment of both the increase and spatial distribution of myocardial perfusion following declamping of an arterial graft.
The laser Doppler technique is used to assess tissue perfusion. Traditionally an integrated, v-weighted (firstorder filter) power spectrum is used to estimate perfusion. In order to be able to obtain selective information about the flow in vessels with different blood cell velocities, higher order filters have been implemented, investigated, and evaluated. Theoretical considerations show that the output of the signal processor will depend on the flow speed, for a given concentration of blood cells, according to Sout}vn where v is the average blood cell speed and n is the spectral filter order. An implementation of filters using zero-, first-, second-, and third-order spectral moments was utilized to experimentally verify the theory by using a laser Doppler perfusion imager. Two different flow models were utilized: A Plexiglas model was used to demonstrate the various signatures of the power spectrum for different flow speeds and filter orders, whereas a Delrin model was used to study the relationship between the flow velocity and the output of the signal processor for the different filters. The results show good agreement with theory and also good reproducibility.
Recordings made on the skin of the wrist area demonstrated that the flow in small veins can be visualized
by the use of higher spectral orders.
Laser Doppler Perfusion Imaging (LDPI) is a method for visualization of tissue blood perfusion. A low power laser beam is used to step-wise scan a tissue area of interest and a perfusion estimate based on the backscattered, partially Doppler broadened, light is generated. Although the basic operating principle of LDPI is the same as that of conventional Laser Doppler Perfusion Monitoring (LDPM), significant differences exist between the implementation of the methods which must be taken into account in order to generate high quality perfusion images. The purpose of this study is to investigate the relevance of a number of LDPI design parameters, such as: (1) The influence of artifact noise when using a continuously moving laser beam instead of a step-wise moving beam to scan the image. (2) The signal processor output's dependency on the distance between the measurement object and the scanner head when using collimated laser light. (3) The speed and mode of the scanning. The results show a substantial rise in the noise level when using a continuously moving beam as opposed to a step-wise. Skin measurements using a collimated laser beam demonstrated an amplification factor dependency on the distance between the skin surface and the scanner head not present when using a divergent laser beam. The scanning speed is limited by the trade-off between the Doppler signal lower cut-off frequency and the image quality.
By the introduction of the laser Doppler perfusion imager (LDPI) the microvascular blood flow in a tissue area can be mapped by sequentially moving a laser beam over the tissue. The measurement is performed without touching the tissue and the captured perfusion values in the peripheral circulation are presented as a color-coded image. In the ordinary LDPI-set-up, 64 X 64 measurement sites cover an area in the range of about 10 - 150 cm2 depending on system settings. With a high resolution modification, recordings can be done on tissue areas as small as 1 cm2. This high resolution option has been assessed in animal models for the mapping of small vessels. To be able to record not only spatial but also temporal perfusion components of tissue blood flow, different local area scans (LAS) have been developed. These include single point recording as well as integration of either 2 X 2, 3 X 3, or 4 X 4 measurement sites. The laser beam is repeatedly moved in a quadratic pattern over the small tissue area of interest and the output value constitutes the average perfusion of all captured values within the actual region. For the evaluation, recordings were performed on healthy volunteers before and after application of a vasodilatating cream on the dorsal side of the hand.
Laser Doppler perfusion monitoring is a method of assessing tissue perfusion based on measurements performed using Doppler broadening of monochromatic light scattered in moving blood cells. Ever since laser Doppler perfusion monitors became available about 15 years ago they have been used in numerous applications in both clinical and laboratory settings. The high spatial resolution has in practice manifested itself as one of the main limitations of the method. The reason for this is the difficulty in attaining reproducible values at successive measurement sites because most skin tissue possesses a substantial variation in blood flow even at adjacent measurement sites. In order to overcome this difficulty the laser Doppler perfusion imager was developed. In this camera-like device, the laser beam successively scans the tissue and the Doppler components of the backscattered light are detected by a remote photodiode. After a scanning procedure is complete, a color-coded perfusion map showing the spatial variation of skin blood flow is displayed on a monitor. The operating principle and early applications of this emerging technology are addressed in further detail.
A laser Doppler perfusion imager has been developed that makes possible mapping of tissue blood flow over surfaces with extensions up to about 12 cm X 12 cm. The He-Ne laser beam scans the tissue under study throughout 4096 measurement sites. A fraction of the backscattered and Doppler broadened light is detected by a photo diode positioned about 20 cm above the tissue surface. After processing, a signal that scales linearly with perfusion is stored in a computer and a color coded image of the spatial tissue perfusion is shown on a monitor. A full format scan is completed in about 4.5 minutes. Algorithms for calculating perfusion profiles and averages as well as substraction of one image from another, form an integral part of the system data analysis software. The perfusion images can also be exported to other software packages for further processing and analysis.
Quasielastic scattering of coherent light in tissue can be utilized to probe the superficial blood flow of the skin and other organs. This fundamental principle has been employed in the construction of laser Doppler tissue perfusion monitors and imagers, intended for experimental and clinical investigations of tissue blood flow. In this paper, the theory of laser Doppler equipment is discussed. The measuring depth can be estimated by Monte Carlo simulation, while the dynamic light scattering (light scattering including Doppler effects) can be modeled by autocorrelation techniques. Based on these techniques estimators for tissue perfusion and tissue blood cell concentration are derived. The operating principle of the laser Doppler monitor is described in detail, with special reference to the design of probes. This monitor is useful for tracking temporal changes in tissue perfusion at a single point. The principle of a recently developed laser Doppler perfusion imager is reviewed. This imager is useful in mapping the spatial variations in tissue perfusion. Different ways of evaluating the performances of the monitor and imager in both mechanical flow simulators and in-vivo studies are discussed. Finally a brief overview is made of some fields of application in which the laser Doppler technique for measurement of tissue perfusion has been successfully used.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.