Of the reported ~20,000 skin disease related deaths in 2013, 30% were attributed to cutaneous inf... more Of the reported ~20,000 skin disease related deaths in 2013, 30% were attributed to cutaneous infections, ulcers, wounds, and burns. Wound healing is currently visually assessed by physicians and commonly requires multiple clinical visits for frequent dressing changes. These are often painful and can further compromise tissue health along with increasing infection risks. Clinical workflows for skin wounds often include oxygenation assessment to help guide the course of treatment. However, the low oxygen perfusion intrinsic to these conditions renders standard-of-care blood oxygen saturation tools inaccurate. Transcutaneous oxygen measurements can overcome this issue, albeit with further complications such as time-consuming bedside calibrations and potentially painful application/removal of electrodes. We present a ‘SMART’ bandage approach that features clinically-validated oxygen sensing molecules embedded within a hydrogel for direct open wound oxygenation measurements. The hydrogel incorporates a phosphorescent red-emitting oxygen-sensing metalloporphyrin covalently attached to a green fluorescent reference dye. As phosphorescence is quenched via collisional energy exchange with molecular oxygen, phosphorescence intensity and lifetime can be used to quantify oxygenation. The emission is bright enough to be visualized by the naked eye, and its intensity was found to increase 10x in oxygen deprived environments (0 mmHg pO2) compared to room air (160 mmHg pO2). Additionally, the hydrogel is capable of swelling up to 400% of its original size in wound exudate while maintaining skin adhesion properties. Along with minimizing the time and materials required for re-dressing, this novel wound dressing has potential for mediating drug delivery and reducing unnecessary patient discomfort.
The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI... more The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI) biomarker and can be used in early diagnosis. As essential primary human immune cells, leukocytes are recruited to injured cerebral sites during TBI response, where they can interact with and potentially bind to TBI biomarkers. To date, no studies have demonstrated ultra‐low GFAP binding enumeration on leukocytes. Herein, a dark‐field imaging technique coupled with computational analysis is introduced to quantify GFAP bound to peripheral blood mononuclear cells (PBMCs). Dark‐field microscopy (DFM) with a custom‐written image acquisition software is developed for rapid 3D PBMC imaging by utilizing specific antiGFAP monoclonal antibody functionalized gold nanoparticles (antiGFAP‐AuNPs) as contrast‐generating probes. Subsequently, the developed algorithm is utilized in processing thousands of acquired images for rapid visualization and enumeration of bound antiGFAP‐AuNP on each leukocyte. The proposed method is demonstrates the specific binding of GFAP to the surface of PBMCs on a healthy donor blood. Thereafter, subpopulations of PBMCs with antiGFAP‐AuNP binding are identified with the assistance of fluorescence imaging and DFM imaging, paving a new way to understanding the relationship between TBI and leukocyte classes. Hence, this study offers a rapid and ultra‐sensitive strategy for biomarker assessment following TBI.
Sensor-integrated wound dressings are emerging tools applicable to a wide variety of medical appl... more Sensor-integrated wound dressings are emerging tools applicable to a wide variety of medical applications from emergency triage to at-home monitoring. Uncomfortable, unnecessary wound dressing changes may be avoided by providing quantitative insight into tissue characteristics related to wound healing such as tissue oxygenation, pH, and exudate/transudate volume. Here, we present a simple cost-effective methodology for quantifying oxygen and pH in a swellable hydrogel dressing using a single photograph. The red and green luminescence of a novel dendritic polyamine Pt-porphyrin and fluorescein conjugate quantitatively responds to oxygen and pH, respectively, and enables robust sensing. The porphyrin conjugate, when combined with a 4-arm star PEG amine polymer, rapidly crosslinks at room temperature with a NHS-PEG crosslinker to form a color-changing hydrogel dressing with tunable swelling capabilities applicable to a variety of wound environments. An inexpensive DSLR camera modified with bandpass filters captures the hydrogel luminescence using simple macroscopic photography, and conversion to HSB colorspace allows for intensity-independent image analysis of the hydrogels' dual modality response. The hydrogel formulation exhibits a robust and validated visible red-orange-green 'traffic light' spectrum in response to oxygen changes, regardless of swelling state, pH, or autofluorescence from skin, thereby enabling clinician friendly naked-eye feedback. This article is protected by copyright. All rights reserved.
Optical Sensors and Sensing Congress 2022 (AIS, LACSEA, Sensors, ES), 2022
We have developed an experimental and algorithmic toolset to obtain reliable transcutaneous oxyge... more We have developed an experimental and algorithmic toolset to obtain reliable transcutaneous oxygenation readings using a lightweight wearable sensor. We present successful clinical results of tissue oxygenation during local and systemic changes in blood flow.
Wearable devices have found widespread applications in recent years as both medical devices as we... more Wearable devices have found widespread applications in recent years as both medical devices as well as consumer electronics for sports and health tracking. A metric of health that is often overlooked in currently available technology is the direct measurement of molecular oxygen in living tissue, a key component in cellular energy production. Here, we report on the development of a wireless wearable prototype for transcutaneous oxygenation monitoring based on quantifying the oxygen-dependent phosphorescence of a metalloporphyrin embedded within a highly breathable oxygen sensing film. The device is completely self-contained, weighs under 30 grams, performs on-board signal analysis, and can communicate with computers or smartphones. The wearable measures tissue oxygenation at the skin surface by detecting the lifetime and intensity of phosphorescence, which undergoes quenching in the presence of oxygen. As well as being insensitive to motion artifacts, it offers robust and reliable measurements even in variable atmospheric conditions related to temperature and humidity. Preliminary in vivo testing in a porcine ischemia model shows that the wearable is highly sensitive to changes in tissue oxygenation in the physiological range upon inducing a decrease in limb perfusion.
Non-fractional lasers used for hair removal penetrate deep into the tissue (~4 mm), and can be re... more Non-fractional lasers used for hair removal penetrate deep into the tissue (~4 mm), and can be repurposed for enhanced thermal delivery using topically applied indocyanine green (ICG), a highly absorptive NIR dye. We demonstrate a new methodology for achieving fractional damage with an 808nm diode laser using a microneedle array injector and ICG impregnated PLGA nanoparticle formulation. A comparison of the effects of injection depth and irradiation dose between free ICG and PLGA@ICG revealed that the nanoparticle formulation effectively concentrates and confines the fluorophore locally at depths of ~3mm and thermal damage is achieved with irradiances as low as 10J/cm2. These improvements in the delivery of ICG subcutaneously in a fractional pattern allow for confined dermal tissue injury using low irradiances, minimizing discoloration of superficial layers of the skin, and significantly enhancing the depth of thermal injury achievable with a wide-area non-fractional laser diode.
The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI... more The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI) biomarker and can be used in early diagnosis. As essential primary human immune cells, leukocytes are recruited to injured cerebral sites during TBI response, where they can interact with and potentially bind to TBI biomarkers. To date, no studies have demonstrated ultra‐low GFAP binding enumeration on leukocytes. Herein, a dark‐field imaging technique coupled with computational analysis is introduced to quantify GFAP bound to peripheral blood mononuclear cells (PBMCs). Dark‐field microscopy (DFM) with a custom‐written image acquisition software is developed for rapid 3D PBMC imaging by utilizing specific antiGFAP monoclonal antibody functionalized gold nanoparticles (antiGFAP‐AuNPs) as contrast‐generating probes. Subsequently, the developed algorithm is utilized in processing thousands of acquired images for rapid visualization and enumeration of bound antiGFAP‐AuNP on each leukocyte....
OSA Optical Sensors and Sensing Congress 2021 (AIS, FTS, HISE, SENSORS, ES), 2021
We have developed wireless wearable devices to monitor tissue oxygenation transcutaneously by det... more We have developed wireless wearable devices to monitor tissue oxygenation transcutaneously by detecting the phosphorescence lifetime and intensity of an oxygen sensing film, paired with machine learning. The devices have been tested in vivo.
Of the reported ~20,000 skin disease related deaths in 2013, 30% were attributed to cutaneous inf... more Of the reported ~20,000 skin disease related deaths in 2013, 30% were attributed to cutaneous infections, ulcers, wounds, and burns. Wound healing is currently visually assessed by physicians and commonly requires multiple clinical visits for frequent dressing changes. These are often painful and can further compromise tissue health along with increasing infection risks. Clinical workflows for skin wounds often include oxygenation assessment to help guide the course of treatment. However, the low oxygen perfusion intrinsic to these conditions renders standard-of-care blood oxygen saturation tools inaccurate. Transcutaneous oxygen measurements can overcome this issue, albeit with further complications such as time-consuming bedside calibrations and potentially painful application/removal of electrodes. We present a ‘SMART’ bandage approach that features clinically-validated oxygen sensing molecules embedded within a hydrogel for direct open wound oxygenation measurements. The hydrogel incorporates a phosphorescent red-emitting oxygen-sensing metalloporphyrin covalently attached to a green fluorescent reference dye. As phosphorescence is quenched via collisional energy exchange with molecular oxygen, phosphorescence intensity and lifetime can be used to quantify oxygenation. The emission is bright enough to be visualized by the naked eye, and its intensity was found to increase 10x in oxygen deprived environments (0 mmHg pO2) compared to room air (160 mmHg pO2). Additionally, the hydrogel is capable of swelling up to 400% of its original size in wound exudate while maintaining skin adhesion properties. Along with minimizing the time and materials required for re-dressing, this novel wound dressing has potential for mediating drug delivery and reducing unnecessary patient discomfort.
The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI... more The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI) biomarker and can be used in early diagnosis. As essential primary human immune cells, leukocytes are recruited to injured cerebral sites during TBI response, where they can interact with and potentially bind to TBI biomarkers. To date, no studies have demonstrated ultra‐low GFAP binding enumeration on leukocytes. Herein, a dark‐field imaging technique coupled with computational analysis is introduced to quantify GFAP bound to peripheral blood mononuclear cells (PBMCs). Dark‐field microscopy (DFM) with a custom‐written image acquisition software is developed for rapid 3D PBMC imaging by utilizing specific antiGFAP monoclonal antibody functionalized gold nanoparticles (antiGFAP‐AuNPs) as contrast‐generating probes. Subsequently, the developed algorithm is utilized in processing thousands of acquired images for rapid visualization and enumeration of bound antiGFAP‐AuNP on each leukocyte. The proposed method is demonstrates the specific binding of GFAP to the surface of PBMCs on a healthy donor blood. Thereafter, subpopulations of PBMCs with antiGFAP‐AuNP binding are identified with the assistance of fluorescence imaging and DFM imaging, paving a new way to understanding the relationship between TBI and leukocyte classes. Hence, this study offers a rapid and ultra‐sensitive strategy for biomarker assessment following TBI.
Sensor-integrated wound dressings are emerging tools applicable to a wide variety of medical appl... more Sensor-integrated wound dressings are emerging tools applicable to a wide variety of medical applications from emergency triage to at-home monitoring. Uncomfortable, unnecessary wound dressing changes may be avoided by providing quantitative insight into tissue characteristics related to wound healing such as tissue oxygenation, pH, and exudate/transudate volume. Here, we present a simple cost-effective methodology for quantifying oxygen and pH in a swellable hydrogel dressing using a single photograph. The red and green luminescence of a novel dendritic polyamine Pt-porphyrin and fluorescein conjugate quantitatively responds to oxygen and pH, respectively, and enables robust sensing. The porphyrin conjugate, when combined with a 4-arm star PEG amine polymer, rapidly crosslinks at room temperature with a NHS-PEG crosslinker to form a color-changing hydrogel dressing with tunable swelling capabilities applicable to a variety of wound environments. An inexpensive DSLR camera modified with bandpass filters captures the hydrogel luminescence using simple macroscopic photography, and conversion to HSB colorspace allows for intensity-independent image analysis of the hydrogels' dual modality response. The hydrogel formulation exhibits a robust and validated visible red-orange-green 'traffic light' spectrum in response to oxygen changes, regardless of swelling state, pH, or autofluorescence from skin, thereby enabling clinician friendly naked-eye feedback. This article is protected by copyright. All rights reserved.
Optical Sensors and Sensing Congress 2022 (AIS, LACSEA, Sensors, ES), 2022
We have developed an experimental and algorithmic toolset to obtain reliable transcutaneous oxyge... more We have developed an experimental and algorithmic toolset to obtain reliable transcutaneous oxygenation readings using a lightweight wearable sensor. We present successful clinical results of tissue oxygenation during local and systemic changes in blood flow.
Wearable devices have found widespread applications in recent years as both medical devices as we... more Wearable devices have found widespread applications in recent years as both medical devices as well as consumer electronics for sports and health tracking. A metric of health that is often overlooked in currently available technology is the direct measurement of molecular oxygen in living tissue, a key component in cellular energy production. Here, we report on the development of a wireless wearable prototype for transcutaneous oxygenation monitoring based on quantifying the oxygen-dependent phosphorescence of a metalloporphyrin embedded within a highly breathable oxygen sensing film. The device is completely self-contained, weighs under 30 grams, performs on-board signal analysis, and can communicate with computers or smartphones. The wearable measures tissue oxygenation at the skin surface by detecting the lifetime and intensity of phosphorescence, which undergoes quenching in the presence of oxygen. As well as being insensitive to motion artifacts, it offers robust and reliable measurements even in variable atmospheric conditions related to temperature and humidity. Preliminary in vivo testing in a porcine ischemia model shows that the wearable is highly sensitive to changes in tissue oxygenation in the physiological range upon inducing a decrease in limb perfusion.
Non-fractional lasers used for hair removal penetrate deep into the tissue (~4 mm), and can be re... more Non-fractional lasers used for hair removal penetrate deep into the tissue (~4 mm), and can be repurposed for enhanced thermal delivery using topically applied indocyanine green (ICG), a highly absorptive NIR dye. We demonstrate a new methodology for achieving fractional damage with an 808nm diode laser using a microneedle array injector and ICG impregnated PLGA nanoparticle formulation. A comparison of the effects of injection depth and irradiation dose between free ICG and PLGA@ICG revealed that the nanoparticle formulation effectively concentrates and confines the fluorophore locally at depths of ~3mm and thermal damage is achieved with irradiances as low as 10J/cm2. These improvements in the delivery of ICG subcutaneously in a fractional pattern allow for confined dermal tissue injury using low irradiances, minimizing discoloration of superficial layers of the skin, and significantly enhancing the depth of thermal injury achievable with a wide-area non-fractional laser diode.
The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI... more The glial fibrillary acidic protein (GFAP) is widely established as a traumatic brain injury (TBI) biomarker and can be used in early diagnosis. As essential primary human immune cells, leukocytes are recruited to injured cerebral sites during TBI response, where they can interact with and potentially bind to TBI biomarkers. To date, no studies have demonstrated ultra‐low GFAP binding enumeration on leukocytes. Herein, a dark‐field imaging technique coupled with computational analysis is introduced to quantify GFAP bound to peripheral blood mononuclear cells (PBMCs). Dark‐field microscopy (DFM) with a custom‐written image acquisition software is developed for rapid 3D PBMC imaging by utilizing specific antiGFAP monoclonal antibody functionalized gold nanoparticles (antiGFAP‐AuNPs) as contrast‐generating probes. Subsequently, the developed algorithm is utilized in processing thousands of acquired images for rapid visualization and enumeration of bound antiGFAP‐AuNP on each leukocyte....
OSA Optical Sensors and Sensing Congress 2021 (AIS, FTS, HISE, SENSORS, ES), 2021
We have developed wireless wearable devices to monitor tissue oxygenation transcutaneously by det... more We have developed wireless wearable devices to monitor tissue oxygenation transcutaneously by detecting the phosphorescence lifetime and intensity of an oxygen sensing film, paired with machine learning. The devices have been tested in vivo.
The controlled assembly of plasmonic nanoparticles by a molecular binding event has emerged as a ... more The controlled assembly of plasmonic nanoparticles by a molecular binding event has emerged as a simple yet sensitive methodology for protein detection. Metallic nanoparticles (NPs) coated with functionalized aptamers can be utilized as biosensors by monitoring changes in particle optical properties, such as the LSPR shift and enhancement of the SERS spectra, in the presence of a target protein. Herein we test this method using two modified aptamers selected for the protein biomarker interleukin 6, an indicator of the dengue fever virus and other diseases including certain types of cancers, diabetes, and even arthritis. IL6 works by inducing an immunological response within the body that can be either anti-inflammatory or pro-inflammatory. The results show that the average hydrodynamic diameter of the NPs as measured by Dynamic Light Scattering was ~42 nm. After conjugation of the aptamers, the peak absorbance of the AgNPs shifted from 404 to 408 nm indicating a surface modification of the NPs due to the presence of the aptamer. Lastly, preliminary results were obtained showing an increase in SERS intensity occurs when the IL-6 protein was introduced to the conjugate solution but the assay will still need to be optimized in order for it to be able to monitor varying concentration changes within and across the desired range.
From the miniaturization of large sample processing machines to the creation of handheld point-of... more From the miniaturization of large sample processing machines to the creation of handheld point-of-care devices, microfluidics has the potential to be a powerful tool in the advancement of diagnostic technologies. Here, we compare different prototyping modalities towards the generation of an inertial microfluidic blood filter: i.e. a 'centrifuge-on-a-chip'. While photolithography is currently the method of choice for soft lithography mold fabrication, offering high design fidelity, we believe simpler methods, such as milling or 3D printing, will soon become equally viable options in the field of microfluidic device fabrication. Three modalities for optofluidic PDMS chip fabrication were compared: micromachining, 3D printing, and SU8 photolithography. The filtration efficiency of the chips were tested with whole blood and compared spectroscopically by monitoring the outlet absorbance at the 540 nm peak intrinsic to oxyhemoglobin at the outlet of each filter chip.
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Papers by Haley Marks