Mira Sibai - Academia.edu (original) (raw)

Papers by Mira Sibai

Frontiers in Physics

Introduction: Fibred-based optical spectroscopy is advantageous over imaging due to its sensitivi... more Introduction: Fibred-based optical spectroscopy is advantageous over imaging due to its sensitivity, practicality and precision, providing point of care diagnosis. The unique advantage is that the sampled volume is well defined by the source-detector geometry and that the functionality of multiple optical techniques can be incorporated into one probe so that more information is gained without extra bulkiness or cost, while also mitigating the limitations of each.Methods: This advantage is utilized here to address the limited accuracy in delineating brain tumors, in situ, by simultaneously characterizing tissue based on the spectral and lifetime properties of five endogenous fluorophores commonly present in brain tissue. A 5-meters multi fibre-optic probe custom-built for neurosurgery guidance with a sterilizable distal end is presented in this paper. It describes its technical features such as architecture, collection efficiency, sensitivity.Results and discussion: The developed pro...

Advanced Biomedical and Clinical Diagnostic and Surgical Guidance Systems XVI, 2018

The goal of fluorescence-guided surgery (FGR) is to provide real-time enhancement of tumors to ma... more The goal of fluorescence-guided surgery (FGR) is to provide real-time enhancement of tumors to maximize safe resection. The optical property mapping ability of spatial frequency domain imaging (SFDI) has enabled quantitative fluorescence imaging (qFI) of protoporphyrin IX (PpIX) in gliomas in the pre-clinical setting. The goal of this study was to evaluate the feasibility of using SFDI to allow for qFI to enhance FGR. Specifically, we modified a benchtop SFDI system to mount directly to a commercial surgical microscope(Zeiss). A commercially available digital light processing module (DLI Austin, TX) was used to modulate light from a xenon arc lamp to illuminate the field. White light excitation and a liquid crystal tunable filter (LCTF Verispec) was used to measure diffuse reflectance at discreet wavelengths from 420 nm to 720 nm on a CMOS camera. An illumination side filter wheel allowed for excitation of PpIX fluorescence at 405 nm and 635 nm and the LCTF measured fluorescence emission at 670 nm and 710 nm. The ability of the clinical microscope to perform optical mapping and qFI was tested with tissue simulating phantoms and live mouse models. The results of these tests showed that SFDI can be implemented in a clinical microscope and the optical mapping and qFI abilities of SFDI may be used to enhance FGR.

Biophotonics: Photonic Solutions for Better Health Care VI, 2018

Intra-operative fluorescence-guided resection (FGR) enables maximum safe resection of glioma by p... more Intra-operative fluorescence-guided resection (FGR) enables maximum safe resection of glioma by providing real-time tumor contrast. In its most widely used form, FGR is mediated by the preferential overproduction of the fluorophore protoporphyrinIX (PpIX) in malignant tissue after an oral dose of its precursor 5-Aminolevulinic Acid (ALA)1. ALA-PpIX-FGR has been shown to significantly increase completeness of tumor resection. However, the subjective visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to delineating only high-grade tumors that display strong fluorescence residing at the tissue surface. We have shown that wide-field quantitative assessment by extracting 2D maps of PpIX concentration in the tissue, [PpIX], significantly improves the accuracy in detecting diffuse tumors, thereby potentially extending FGR to patients with low-grade tumors. In this approach, hyperspectral fluorescence imaging is coupled to a custom-built spatial frequency domain imaging (SFDI) system. SFDI enables the recovery of tissue optical properties maps, μ_a and μ_s^'. These are used to correct the fluorescence images. The corrected hyperspectral fluorescence images are then spectrally unmixed to separate true PpIX fluorescence from that of its photoproducts and from autofluorescence. Quantitative fluorescence imaging was validated against the clinically used spectroscopic probe by comparing the recovered optical properties and [PpIX] in vivo of a rat brain tumor model. This quantitative approach was also applied to a near infrared fluorophore ZW-800 on tissue-simulating phantoms. ALA-PpIX-FGR, as it is currently implemented, is inaccessible to infiltrative residuals lying beyond the resection cavity because of the limited penetration depth of the blue excitation light used. This is problematic as these infiltrative tumors are the main cause of reccurence. Enhanced sub-surface tumor detection was shown feasible by exciting PpIX’s secondary absorption peak of 635 nm intra-operatively on patients with various intra-cranial pathologies. However, resolving strong fluorescence of a deep-seated tumor from weak fluorescence of a shallow tumor was not possible. That is because the detected fluorescence intensity is heavily dependent on fluorophore concentration, depth, and fluorophore distribution, while also being convolved with tissue turbidity. The aim of this work, therefore, is to extend quantitative ALA-PpIX-FGR to identify sub-surface tumors by resolving tumor depth from fluorophore concentration. This should assist the surgeon in making an informed decision as for whether to further resect or not. A new quantitative depth imaging method was developed by exploiting SFDI’s depth-encoding capabilities in fluorescence mode. The result is a series of spatially modulated fluorescence images, where the modulation amplitude decays with increasing spatial frequency at a rate dependent on fluorophore depth. After recovering depth, a diffusion-based fluorescent light transport model is applied to extract fluorophore concentration. The algorithm was validated using tissue-simulating phantoms and an ex vivo tissue model indicating that the maximum depth recovered is highly dependent on fluorophore concentration as well as on tissue turbidity. For the [PpIX] and optical property maps relevant for glioma tissue, our quantitative depth fluorescence technique can predict depths up to 9 mm ± 0.4 mm, while recovering [PpIX] with an accuracy of 15% for concentrations as low as 2.5 µg/ml.

The extent of resection is now considered a significant prognostic factor; yet, safe complete res... more The extent of resection is now considered a significant prognostic factor; yet, safe complete resection is achieved in only 30% glioma patients. The challenge of surgery arises from the diffusive nature of gliomas known to infiltrate normal parenchyma beyond the resection cavity, which are left undetected. To better visualize glioma, intra-operative fluorescence-guided resection (FGR) has been a practical solution, providing real-time tumor contrast. In its most widely used form, FGR is mediated by the preferential overproduction of the fluorophore protoporphyrin IX (PpIX) in malignant tissue after an oral dose of its precursor 5-Aminolevulinic Acid (ALA). ALA-PpIX-FGR has been shown to significantly increase extent of resection. However, the visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to delineating only high-grade tumors that display strong fluorescence. To this end, the work described in this thesis outlines the development of i...

Biomedical Optics Express, 2020

Fluorescence-guided surgery (FGS) enhances intraoperative visualization of tumors to maximize saf... more Fluorescence-guided surgery (FGS) enhances intraoperative visualization of tumors to maximize safe resection, and quantitative fluorescence imaging (qFI) of protoporphyrin IX (PpIX) has provided additional information for guidance during intracranial tumor surgery. Previous developments in fluorescence quantification have demonstrated that the depth of fluorescence signals can be estimated given known optical properties in a lab setting, and now with the work described here that these optical properties can be determined in vivo in human brain tissue in the operating room (OR) during tumor resection procedures. More specifically, we report the first depth estimation of subsurface tumor intraoperatively, achieved with the combination of spatial frequency domain imaging (SFDI) for optical property measurement and red-light excitation of PpIX. We modified a commercial surgical microscope (Zeiss) with a digital light processing module (DLI Austin, TX) to modulate light from a xenon arc ...

Journal of Biophotonics, 2019

Two photon fluorescence images (a,b,c) and its corresponding histological images (d,e,f) and the ... more Two photon fluorescence images (a,b,c) and its corresponding histological images (d,e,f) and the autofluorescence spectra under 405 nm, 810 nm and 890 nm excitation (g,h,i) of different human brain tissues, from three different types : control(a,d,g), low grade glioma tumor(b,e,h) and glioblastoma tumor(c,f,i).

Journal of Biophotonics, 2019

Journal of Biophotonics, 2018

To complement a project towards label-free optical biopsy and enhanced resection which the overal... more To complement a project towards label-free optical biopsy and enhanced resection which the overall goal is to develop a multimodal non-linear endomicroscope, this multimodal approach aims to enhance the accuracy in classifying brain tissue into solid tumor, infiltration, and normal tissue intraoperatively. Multiple optical measurements based on one and two-photon spectral and lifetime autofluorescence, including second harmonic generation imaging were acquired. As a prerequisite, studying the effect of the time of measurement post-excision on tissue's spectral/lifetime fluorescence properties was warranted, so spectral and lifetime fluorescence of fresh brain tissues were measured using a point-based linear endoscope. Additionally, a comparative study on tissue's optical properties obtained by multimodal non-linear optical imaging microscope from fresh and fixed tissue was necessary to test whether clinical validation of the non-linear endomicroscope is feasible by extracting optical signatures from fixed tissue rather than from freshly excised samples. The former is generally chosen for convenience. Results of this study suggest that an hour is necessary post-excision to have consistent fluorescence intensities\lifetimes. The fresh vs. fixed tissue study indicates that while all optical signals differ after fixation, the characteristic features extracted from one and twophoton excitation still discriminate normal brain cortical tissue, glioblastoma, and metastases. Picture: Two photon fluorescence images of fresh (a,b,c) and fixed (d,e,f) human brain tissues, from three different types : control(a,d), GBM tumor(b,e) and metastasis tumor(c,f). K E Y W O R D S Fluorescence, multiphoton microscopy, fluorescence lifetime imaging, spectroscopy, fresh and fixed human brain tissues.

Scientific reports, Jan 24, 2018

Accurate intraoperative tumour margin assessment is a major challenge in neurooncology, where spa... more Accurate intraoperative tumour margin assessment is a major challenge in neurooncology, where sparse tumours beyond the bulk tumour are left undetected under conventional resection. Non-linear optical imaging can diagnose tissue at the sub-micron level and provide functional label-free histopathology in vivo. For this reason, a non-linear endomicroscope is being developed to characterize brain tissue intraoperatively based on multiple endogenous optical contrasts such as spectrally- and temporally-resolved fluorescence. To produce highly sensitive optical signatures that are specific to a given tissue type, short femtosecond pulsed lasers are required for efficient two-photon excitation. Yet, the potential of causing bio-damage has not been studied on neuronal tissue. Therefore, as a prerequisite to clinically testing the non-linear endomicroscope in vivo, the effect of short laser pulse durations (40-340 fs) on ex vivo brain tissue was investigated by monitoring the intensity, the ...

Journal of Biomedical Optics, 2018

Mapping the optical absorption and scattering properties of tissues using spatial frequency-domai... more Mapping the optical absorption and scattering properties of tissues using spatial frequency-domain imaging (SFDI) enhances quantitative fluorescence imaging of protoporphyrin IX (PpIX) in gliomas in the preclinical setting. The feasibility of using SFDI in the operating room was investigated here. A benchtop SFDI system was modified to mount directly to a commercial operating microscope. A digital light processing module imposed a selectable spatial light pattern from a broad-band xenon arc lamp to illuminate the surgical field. White light excitation and a liquid crystal-tunable filter allowed the diffuse reflectance images to be recorded at discrete wavelengths from 450 to 720 nm on a sCMOS camera. The performance was first tested in tissue-simulating phantoms, and data were then acquired intraoperatively during brain tumor resection surgery. The optical absorption and transport scattering coefficients could be estimated with average errors of 3.2% and 4.5% for the benchtop and clinical systems, respectively, with spatial resolution of better than 0.7 mm. These findings suggest that SFDI can be implemented in a clinically relevant configuration to achieve accurate mapping of the optical properties in the surgical field that can then be applied to achieve quantitative imaging of the fluorophore.

Journal of biomedical optics, Jul 1, 2017

5-Aminolevelunic acid-induced protoporphyrin IX (PpIX) fluorescence-guided resection (FGR) enable... more 5-Aminolevelunic acid-induced protoporphyrin IX (PpIX) fluorescence-guided resection (FGR) enables maximum safe resection of glioma by providing real-time tumor contrast. However, the subjective visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to reliably delineating only high-grade tumors that display strong fluorescence. We have previously shown, using a fiber-optic probe, that quantitative assessment using noninvasive point spectroscopic measurements of the absolute PpIX concentration in tissue further improves the accuracy of FGR, extending it to surgically curable low-grade glioma. More recently, we have shown that implementing spatial frequency domain imaging with a fluorescent-light transport model enables recovery of two-dimensional images of [PpIX], alleviating the need for time-consuming point sampling of the brain surface. We present first results of this technique modified for <italic<in vivo</italic< imaging on ...

Clinical and Translational Neurophotonics; Neural Imaging and Sensing; and Optogenetics and Optical Manipulation, 2016

Cancer tissue often remains after brain tumor resection due to the inability to detect the full e... more Cancer tissue often remains after brain tumor resection due to the inability to detect the full extent of cancer during surgery, particularly near tumor boundaries. Commercial systems are available for intra-operative real-time aminolevulenic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence imaging. These are standard white-light neurosurgical microscopes adapted with optical components for fluorescence excitation and detection. However, these instruments lack sensitivity and specificity, which limits the ability to detect low levels of PpIX and distinguish it from tissue auto-fluorescence. Current systems also cannot provide repeatable and un-biased quantitative fluorophore concentration values because of the unknown and highly variable light attenuation by tissue. We present a highly sensitive spectroscopic fluorescence imaging system that is seamlessly integrated onto a neurosurgical microscope. Hardware and software were developed to achieve through-microscope spatially-modulated illumination for 3D profilometry and to use this information to extract tissue optical properties to correct for the effects of tissue light attenuation. This gives pixel-by-pixel quantified fluorescence values and improves detection of low PpIX concentrations. This is achieved using a high-sensitivity Electron Multiplying Charge Coupled Device (EMCCD) with a Liquid Crystal Tunable Filter (LCTF) whereby spectral bands are acquired sequentially; and a snapshot camera system with simultaneous acquisition of all bands is used for profilometry and optical property recovery. Sensitivity and specificity to PpIX is demonstrated using brain tissue phantoms and intraoperative human data acquired in an on-going clinical study using PpIX fluorescence to guide glioma resection.

Biomedical optics express, 2015

Intraoperative 5- aminolevulinic acid induced-Protoporphyrin IX (PpIX) fluorescence guidance enab... more Intraoperative 5- aminolevulinic acid induced-Protoporphyrin IX (PpIX) fluorescence guidance enables maximum safe resection of glioblastomas by providing surgeons with real-time tumor optical contrast. However, visual assessment of PpIX fluorescence is subjective and limited by the distorting effects of light attenuation and tissue autofluorescence. We have previously shown that non-invasive point measurements of absolute PpIX concentration identifies residual tumor that is otherwise non-detectable. Here, we extend this approach to wide-field quantitative fluorescence imaging by implementing spatial frequency domain imaging to recover tissue optical properties across the field-of-view in phantoms and ex vivo tissue.

Biomedical optics express, 2015

In glioma surgery, Protoporphyrin IX (PpIX) fluorescence may identify residual tumor that could b... more In glioma surgery, Protoporphyrin IX (PpIX) fluorescence may identify residual tumor that could be resected while minimizing damage to normal brain. We demonstrate that improved sensitivity for wide-field spectroscopic fluorescence imaging is achieved with minimal disruption to the neurosurgical workflow using an electron-multiplying charge-coupled device (EMCCD) relative to a state-of-the-art CMOS system. In phantom experiments the EMCCD system can detect at least two orders-of-magnitude lower PpIX. Ex vivo tissue imaging on a rat glioma model demonstrates improved fluorescence contrast compared with neurosurgical fluorescence microscope technology, and the fluorescence detection is confirmed with measurements from a clinically-validated spectroscopic probe. Greater PpIX sensitivity in wide-field fluorescence imaging may improve the residual tumor detection during surgery with consequent impact on survival.

We present an imaging system for which optical components are seamlessly integrated to a commerci... more We present an imaging system for which optical components are seamlessly integrated to a commercial neurosurgical microscope. The system integrates: (1) A broad-beam excitation system with wavelengths between 390nm and 635nm; (2) A spatially-modulated digital light projector that projects patterns of varying spatial frequencies and orientations on the surgical cavity, (3) a dual camera system connected through a coherent bundle to the microscope that can provide structural detection of wide-field hyper-spectral reflectance and fluorescence images. The technique has been developed for fluorescence-guided brain cancer surgery but it can be applied to other organs when coupled with the right molecular marker(s).

Molecular-Guided Surgery: Molecules, Devices, and Applications, 2015

Intraoperative fluorescence guidance enables maximum safe resection of, for example, glioblastoma... more Intraoperative fluorescence guidance enables maximum safe resection of, for example, glioblastomas by providing surgeons with real-time tumor optical contrast. Specifically, 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence guided resection can improve surgical outcomes by better defining tumor margins and identifying satellite tumor foci. However, visual assessment of PpIX fluorescence is subjective and limited by the distorting effects of light attenuation (absorption and scattering) by tissue and background tissue autofluorescence. We have previously shown, using a point fluorescence-reflectance fiberoptic probe, that non-invasive measurement of the absolute PpIX concentration, [PpIX], further improves sensitivity and specificity, leading to the demonstration that the technique can also detect low-grade gliomas as well as otherwise undetectable residual tumor foci in high-grade disease. Here, we extend this approach to wide-field quantitative fluorescence imaging (qFI) by implementing spatial frequency domain imaging (SFDI) to recover the tissue optical absorption and transport scattering coefficients across the field of view. We report on the performance of this approach to determine the intrinsic fluorescence intensity in tissue-simulating phantoms in both the fully diffusive (i.e. scatter-dominated) and sub-diffusive (low transport albedo) regimes, for which higher spatial frequencies are used. The performance of qFI is compared to a Born- normalization correction scheme, as well as to the values obtained using the fiberoptic probe on homogeneous tissue phantoms containing PpIX.

I hereby declare that I am the sole author of this thesis. I authorize Ryerson University to lend... more I hereby declare that I am the sole author of this thesis. I authorize Ryerson University to lend this thesis to other institutions or individuals for the purpose of scholarly research.

Frontiers in Physics

Introduction: Fibred-based optical spectroscopy is advantageous over imaging due to its sensitivi... more Introduction: Fibred-based optical spectroscopy is advantageous over imaging due to its sensitivity, practicality and precision, providing point of care diagnosis. The unique advantage is that the sampled volume is well defined by the source-detector geometry and that the functionality of multiple optical techniques can be incorporated into one probe so that more information is gained without extra bulkiness or cost, while also mitigating the limitations of each.Methods: This advantage is utilized here to address the limited accuracy in delineating brain tumors, in situ, by simultaneously characterizing tissue based on the spectral and lifetime properties of five endogenous fluorophores commonly present in brain tissue. A 5-meters multi fibre-optic probe custom-built for neurosurgery guidance with a sterilizable distal end is presented in this paper. It describes its technical features such as architecture, collection efficiency, sensitivity.Results and discussion: The developed pro...

Advanced Biomedical and Clinical Diagnostic and Surgical Guidance Systems XVI, 2018

The goal of fluorescence-guided surgery (FGR) is to provide real-time enhancement of tumors to ma... more The goal of fluorescence-guided surgery (FGR) is to provide real-time enhancement of tumors to maximize safe resection. The optical property mapping ability of spatial frequency domain imaging (SFDI) has enabled quantitative fluorescence imaging (qFI) of protoporphyrin IX (PpIX) in gliomas in the pre-clinical setting. The goal of this study was to evaluate the feasibility of using SFDI to allow for qFI to enhance FGR. Specifically, we modified a benchtop SFDI system to mount directly to a commercial surgical microscope(Zeiss). A commercially available digital light processing module (DLI Austin, TX) was used to modulate light from a xenon arc lamp to illuminate the field. White light excitation and a liquid crystal tunable filter (LCTF Verispec) was used to measure diffuse reflectance at discreet wavelengths from 420 nm to 720 nm on a CMOS camera. An illumination side filter wheel allowed for excitation of PpIX fluorescence at 405 nm and 635 nm and the LCTF measured fluorescence emission at 670 nm and 710 nm. The ability of the clinical microscope to perform optical mapping and qFI was tested with tissue simulating phantoms and live mouse models. The results of these tests showed that SFDI can be implemented in a clinical microscope and the optical mapping and qFI abilities of SFDI may be used to enhance FGR.

Biophotonics: Photonic Solutions for Better Health Care VI, 2018

Intra-operative fluorescence-guided resection (FGR) enables maximum safe resection of glioma by p... more Intra-operative fluorescence-guided resection (FGR) enables maximum safe resection of glioma by providing real-time tumor contrast. In its most widely used form, FGR is mediated by the preferential overproduction of the fluorophore protoporphyrinIX (PpIX) in malignant tissue after an oral dose of its precursor 5-Aminolevulinic Acid (ALA)1. ALA-PpIX-FGR has been shown to significantly increase completeness of tumor resection. However, the subjective visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to delineating only high-grade tumors that display strong fluorescence residing at the tissue surface. We have shown that wide-field quantitative assessment by extracting 2D maps of PpIX concentration in the tissue, [PpIX], significantly improves the accuracy in detecting diffuse tumors, thereby potentially extending FGR to patients with low-grade tumors. In this approach, hyperspectral fluorescence imaging is coupled to a custom-built spatial frequency domain imaging (SFDI) system. SFDI enables the recovery of tissue optical properties maps, μ_a and μ_s^'. These are used to correct the fluorescence images. The corrected hyperspectral fluorescence images are then spectrally unmixed to separate true PpIX fluorescence from that of its photoproducts and from autofluorescence. Quantitative fluorescence imaging was validated against the clinically used spectroscopic probe by comparing the recovered optical properties and [PpIX] in vivo of a rat brain tumor model. This quantitative approach was also applied to a near infrared fluorophore ZW-800 on tissue-simulating phantoms. ALA-PpIX-FGR, as it is currently implemented, is inaccessible to infiltrative residuals lying beyond the resection cavity because of the limited penetration depth of the blue excitation light used. This is problematic as these infiltrative tumors are the main cause of reccurence. Enhanced sub-surface tumor detection was shown feasible by exciting PpIX’s secondary absorption peak of 635 nm intra-operatively on patients with various intra-cranial pathologies. However, resolving strong fluorescence of a deep-seated tumor from weak fluorescence of a shallow tumor was not possible. That is because the detected fluorescence intensity is heavily dependent on fluorophore concentration, depth, and fluorophore distribution, while also being convolved with tissue turbidity. The aim of this work, therefore, is to extend quantitative ALA-PpIX-FGR to identify sub-surface tumors by resolving tumor depth from fluorophore concentration. This should assist the surgeon in making an informed decision as for whether to further resect or not. A new quantitative depth imaging method was developed by exploiting SFDI’s depth-encoding capabilities in fluorescence mode. The result is a series of spatially modulated fluorescence images, where the modulation amplitude decays with increasing spatial frequency at a rate dependent on fluorophore depth. After recovering depth, a diffusion-based fluorescent light transport model is applied to extract fluorophore concentration. The algorithm was validated using tissue-simulating phantoms and an ex vivo tissue model indicating that the maximum depth recovered is highly dependent on fluorophore concentration as well as on tissue turbidity. For the [PpIX] and optical property maps relevant for glioma tissue, our quantitative depth fluorescence technique can predict depths up to 9 mm ± 0.4 mm, while recovering [PpIX] with an accuracy of 15% for concentrations as low as 2.5 µg/ml.

The extent of resection is now considered a significant prognostic factor; yet, safe complete res... more The extent of resection is now considered a significant prognostic factor; yet, safe complete resection is achieved in only 30% glioma patients. The challenge of surgery arises from the diffusive nature of gliomas known to infiltrate normal parenchyma beyond the resection cavity, which are left undetected. To better visualize glioma, intra-operative fluorescence-guided resection (FGR) has been a practical solution, providing real-time tumor contrast. In its most widely used form, FGR is mediated by the preferential overproduction of the fluorophore protoporphyrin IX (PpIX) in malignant tissue after an oral dose of its precursor 5-Aminolevulinic Acid (ALA). ALA-PpIX-FGR has been shown to significantly increase extent of resection. However, the visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to delineating only high-grade tumors that display strong fluorescence. To this end, the work described in this thesis outlines the development of i...

Biomedical Optics Express, 2020

Fluorescence-guided surgery (FGS) enhances intraoperative visualization of tumors to maximize saf... more Fluorescence-guided surgery (FGS) enhances intraoperative visualization of tumors to maximize safe resection, and quantitative fluorescence imaging (qFI) of protoporphyrin IX (PpIX) has provided additional information for guidance during intracranial tumor surgery. Previous developments in fluorescence quantification have demonstrated that the depth of fluorescence signals can be estimated given known optical properties in a lab setting, and now with the work described here that these optical properties can be determined in vivo in human brain tissue in the operating room (OR) during tumor resection procedures. More specifically, we report the first depth estimation of subsurface tumor intraoperatively, achieved with the combination of spatial frequency domain imaging (SFDI) for optical property measurement and red-light excitation of PpIX. We modified a commercial surgical microscope (Zeiss) with a digital light processing module (DLI Austin, TX) to modulate light from a xenon arc ...

Journal of Biophotonics, 2019

Two photon fluorescence images (a,b,c) and its corresponding histological images (d,e,f) and the ... more Two photon fluorescence images (a,b,c) and its corresponding histological images (d,e,f) and the autofluorescence spectra under 405 nm, 810 nm and 890 nm excitation (g,h,i) of different human brain tissues, from three different types : control(a,d,g), low grade glioma tumor(b,e,h) and glioblastoma tumor(c,f,i).

Journal of Biophotonics, 2019

Journal of Biophotonics, 2018

To complement a project towards label-free optical biopsy and enhanced resection which the overal... more To complement a project towards label-free optical biopsy and enhanced resection which the overall goal is to develop a multimodal non-linear endomicroscope, this multimodal approach aims to enhance the accuracy in classifying brain tissue into solid tumor, infiltration, and normal tissue intraoperatively. Multiple optical measurements based on one and two-photon spectral and lifetime autofluorescence, including second harmonic generation imaging were acquired. As a prerequisite, studying the effect of the time of measurement post-excision on tissue's spectral/lifetime fluorescence properties was warranted, so spectral and lifetime fluorescence of fresh brain tissues were measured using a point-based linear endoscope. Additionally, a comparative study on tissue's optical properties obtained by multimodal non-linear optical imaging microscope from fresh and fixed tissue was necessary to test whether clinical validation of the non-linear endomicroscope is feasible by extracting optical signatures from fixed tissue rather than from freshly excised samples. The former is generally chosen for convenience. Results of this study suggest that an hour is necessary post-excision to have consistent fluorescence intensities\lifetimes. The fresh vs. fixed tissue study indicates that while all optical signals differ after fixation, the characteristic features extracted from one and twophoton excitation still discriminate normal brain cortical tissue, glioblastoma, and metastases. Picture: Two photon fluorescence images of fresh (a,b,c) and fixed (d,e,f) human brain tissues, from three different types : control(a,d), GBM tumor(b,e) and metastasis tumor(c,f). K E Y W O R D S Fluorescence, multiphoton microscopy, fluorescence lifetime imaging, spectroscopy, fresh and fixed human brain tissues.

Scientific reports, Jan 24, 2018

Accurate intraoperative tumour margin assessment is a major challenge in neurooncology, where spa... more Accurate intraoperative tumour margin assessment is a major challenge in neurooncology, where sparse tumours beyond the bulk tumour are left undetected under conventional resection. Non-linear optical imaging can diagnose tissue at the sub-micron level and provide functional label-free histopathology in vivo. For this reason, a non-linear endomicroscope is being developed to characterize brain tissue intraoperatively based on multiple endogenous optical contrasts such as spectrally- and temporally-resolved fluorescence. To produce highly sensitive optical signatures that are specific to a given tissue type, short femtosecond pulsed lasers are required for efficient two-photon excitation. Yet, the potential of causing bio-damage has not been studied on neuronal tissue. Therefore, as a prerequisite to clinically testing the non-linear endomicroscope in vivo, the effect of short laser pulse durations (40-340 fs) on ex vivo brain tissue was investigated by monitoring the intensity, the ...

Journal of Biomedical Optics, 2018

Mapping the optical absorption and scattering properties of tissues using spatial frequency-domai... more Mapping the optical absorption and scattering properties of tissues using spatial frequency-domain imaging (SFDI) enhances quantitative fluorescence imaging of protoporphyrin IX (PpIX) in gliomas in the preclinical setting. The feasibility of using SFDI in the operating room was investigated here. A benchtop SFDI system was modified to mount directly to a commercial operating microscope. A digital light processing module imposed a selectable spatial light pattern from a broad-band xenon arc lamp to illuminate the surgical field. White light excitation and a liquid crystal-tunable filter allowed the diffuse reflectance images to be recorded at discrete wavelengths from 450 to 720 nm on a sCMOS camera. The performance was first tested in tissue-simulating phantoms, and data were then acquired intraoperatively during brain tumor resection surgery. The optical absorption and transport scattering coefficients could be estimated with average errors of 3.2% and 4.5% for the benchtop and clinical systems, respectively, with spatial resolution of better than 0.7 mm. These findings suggest that SFDI can be implemented in a clinically relevant configuration to achieve accurate mapping of the optical properties in the surgical field that can then be applied to achieve quantitative imaging of the fluorophore.

Journal of biomedical optics, Jul 1, 2017

5-Aminolevelunic acid-induced protoporphyrin IX (PpIX) fluorescence-guided resection (FGR) enable... more 5-Aminolevelunic acid-induced protoporphyrin IX (PpIX) fluorescence-guided resection (FGR) enables maximum safe resection of glioma by providing real-time tumor contrast. However, the subjective visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to reliably delineating only high-grade tumors that display strong fluorescence. We have previously shown, using a fiber-optic probe, that quantitative assessment using noninvasive point spectroscopic measurements of the absolute PpIX concentration in tissue further improves the accuracy of FGR, extending it to surgically curable low-grade glioma. More recently, we have shown that implementing spatial frequency domain imaging with a fluorescent-light transport model enables recovery of two-dimensional images of [PpIX], alleviating the need for time-consuming point sampling of the brain surface. We present first results of this technique modified for <italic<in vivo</italic< imaging on ...

Clinical and Translational Neurophotonics; Neural Imaging and Sensing; and Optogenetics and Optical Manipulation, 2016

Cancer tissue often remains after brain tumor resection due to the inability to detect the full e... more Cancer tissue often remains after brain tumor resection due to the inability to detect the full extent of cancer during surgery, particularly near tumor boundaries. Commercial systems are available for intra-operative real-time aminolevulenic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence imaging. These are standard white-light neurosurgical microscopes adapted with optical components for fluorescence excitation and detection. However, these instruments lack sensitivity and specificity, which limits the ability to detect low levels of PpIX and distinguish it from tissue auto-fluorescence. Current systems also cannot provide repeatable and un-biased quantitative fluorophore concentration values because of the unknown and highly variable light attenuation by tissue. We present a highly sensitive spectroscopic fluorescence imaging system that is seamlessly integrated onto a neurosurgical microscope. Hardware and software were developed to achieve through-microscope spatially-modulated illumination for 3D profilometry and to use this information to extract tissue optical properties to correct for the effects of tissue light attenuation. This gives pixel-by-pixel quantified fluorescence values and improves detection of low PpIX concentrations. This is achieved using a high-sensitivity Electron Multiplying Charge Coupled Device (EMCCD) with a Liquid Crystal Tunable Filter (LCTF) whereby spectral bands are acquired sequentially; and a snapshot camera system with simultaneous acquisition of all bands is used for profilometry and optical property recovery. Sensitivity and specificity to PpIX is demonstrated using brain tissue phantoms and intraoperative human data acquired in an on-going clinical study using PpIX fluorescence to guide glioma resection.

Biomedical optics express, 2015

Intraoperative 5- aminolevulinic acid induced-Protoporphyrin IX (PpIX) fluorescence guidance enab... more Intraoperative 5- aminolevulinic acid induced-Protoporphyrin IX (PpIX) fluorescence guidance enables maximum safe resection of glioblastomas by providing surgeons with real-time tumor optical contrast. However, visual assessment of PpIX fluorescence is subjective and limited by the distorting effects of light attenuation and tissue autofluorescence. We have previously shown that non-invasive point measurements of absolute PpIX concentration identifies residual tumor that is otherwise non-detectable. Here, we extend this approach to wide-field quantitative fluorescence imaging by implementing spatial frequency domain imaging to recover tissue optical properties across the field-of-view in phantoms and ex vivo tissue.

Biomedical optics express, 2015

In glioma surgery, Protoporphyrin IX (PpIX) fluorescence may identify residual tumor that could b... more In glioma surgery, Protoporphyrin IX (PpIX) fluorescence may identify residual tumor that could be resected while minimizing damage to normal brain. We demonstrate that improved sensitivity for wide-field spectroscopic fluorescence imaging is achieved with minimal disruption to the neurosurgical workflow using an electron-multiplying charge-coupled device (EMCCD) relative to a state-of-the-art CMOS system. In phantom experiments the EMCCD system can detect at least two orders-of-magnitude lower PpIX. Ex vivo tissue imaging on a rat glioma model demonstrates improved fluorescence contrast compared with neurosurgical fluorescence microscope technology, and the fluorescence detection is confirmed with measurements from a clinically-validated spectroscopic probe. Greater PpIX sensitivity in wide-field fluorescence imaging may improve the residual tumor detection during surgery with consequent impact on survival.

We present an imaging system for which optical components are seamlessly integrated to a commerci... more We present an imaging system for which optical components are seamlessly integrated to a commercial neurosurgical microscope. The system integrates: (1) A broad-beam excitation system with wavelengths between 390nm and 635nm; (2) A spatially-modulated digital light projector that projects patterns of varying spatial frequencies and orientations on the surgical cavity, (3) a dual camera system connected through a coherent bundle to the microscope that can provide structural detection of wide-field hyper-spectral reflectance and fluorescence images. The technique has been developed for fluorescence-guided brain cancer surgery but it can be applied to other organs when coupled with the right molecular marker(s).

Molecular-Guided Surgery: Molecules, Devices, and Applications, 2015

Intraoperative fluorescence guidance enables maximum safe resection of, for example, glioblastoma... more Intraoperative fluorescence guidance enables maximum safe resection of, for example, glioblastomas by providing surgeons with real-time tumor optical contrast. Specifically, 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence guided resection can improve surgical outcomes by better defining tumor margins and identifying satellite tumor foci. However, visual assessment of PpIX fluorescence is subjective and limited by the distorting effects of light attenuation (absorption and scattering) by tissue and background tissue autofluorescence. We have previously shown, using a point fluorescence-reflectance fiberoptic probe, that non-invasive measurement of the absolute PpIX concentration, [PpIX], further improves sensitivity and specificity, leading to the demonstration that the technique can also detect low-grade gliomas as well as otherwise undetectable residual tumor foci in high-grade disease. Here, we extend this approach to wide-field quantitative fluorescence imaging (qFI) by implementing spatial frequency domain imaging (SFDI) to recover the tissue optical absorption and transport scattering coefficients across the field of view. We report on the performance of this approach to determine the intrinsic fluorescence intensity in tissue-simulating phantoms in both the fully diffusive (i.e. scatter-dominated) and sub-diffusive (low transport albedo) regimes, for which higher spatial frequencies are used. The performance of qFI is compared to a Born- normalization correction scheme, as well as to the values obtained using the fiberoptic probe on homogeneous tissue phantoms containing PpIX.

I hereby declare that I am the sole author of this thesis. I authorize Ryerson University to lend... more I hereby declare that I am the sole author of this thesis. I authorize Ryerson University to lend this thesis to other institutions or individuals for the purpose of scholarly research.