A handheld device for intra-cavity and ex vivo fluorescence imaging of breast conserving surgery margins with 5-aminolevulinic acid.

BACKGROUND: Visualization of cancer during breast conserving surgery (BCS) remains challenging; the BCS reoperation rate is reported to be 20-70% of patients. An urgent clinical need exists for real-time intraoperative visualization of breast carcinomas during BCS. We previously demonstrated the ability of a prototype imaging device to identify breast carcinoma in excised surgical specimens following 5-aminolevulinic acid (5-ALA) administration. However, this prototype device was not designed to image the surgical cavity for remaining carcinoma after the excised lumpectomy specimen is removed. A new handheld fluorescence (FL) imaging prototype device, designed to image both excised specimens and within the surgical cavity, was assessed in a clinical trial to evaluate its clinical utility for first-in-human, real-time intraoperative imaging during index BCS. RESULTS: The imaging device combines consumer-grade imaging sensory technology with miniature light-emitting diodes (LEDs) and multiband optical filtering to capture high-resolution white light (WL) and FL digital images and videos. The technology allows for visualization of protoporphyrin IX (PpIX), which fluoresces red when excited by violet-blue light. To date, n = 17 patients have received 20 mg kg bodyweight (BW) 5-ALA orally 2-4 h before imaging to facilitate the accumulation of PpIX within tumour cells. Tissue types were identified based on their colour appearance. Breast tumours in sectioned lumpectomies appeared red, which contrasted against the green connective tissues and orange-brown adipose tissues. In addition, ductal carcinoma in situ (DCIS) that was missed during intraoperative standard of care was identified at the surgical margin at <1 mm depth. In addition, artifacts due to the surgical drape, illumination, and blood within the surgical cavity were discovered. CONCLUSIONS: This study has demonstrated the detection of a grossly occult positive margin intraoperatively. Artifacts from imaging within the surgical cavity have been identified, and potential mitigations have been proposed. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01837225 (Trial start date is September 2010. It was registered to ClinicalTrials.gov retrospectively on April 23, 2013, then later updated on April 9, 2020, to reflect the introduction of the new imaging device.).

Customizable optical tissue phantom platform for characterization of fluorescence imaging device sensitivity.

Significance: Optical tissue phantoms serve as inanimate and stable reference materials used to calibrate, characterize, standardize, and test biomedical imaging instruments. Although various types of solid tissue phantoms have been described in the literature, current phantom models are limited in that they do not have a depth feature that can be adjusted in real-time, they cannot be adapted to other applications, and their fabrication can be laborious and costly. Aim: Our goal was to develop an optical phantom that could assess the imaging performance of fluorescence imaging devices and be customizable for different applications. Approach: We developed a phantom with three distinct components, each of which can be customized. Results: We present a method for fabricating a solid optical tissue that contains (1) an adjustable depth capability using thin film phantoms, (2) a refillable chip loaded with fluorophores of the user's choice in various desired quantities, and (3) phantom materials representative of different tissue types. Conclusions: This article describes the development of phantom models that are customizable, adaptable, and easy to design and fabricate.

Quantitative intravital imaging for real-time monitoring of pancreatic tumor cell hypoxia and stroma in an orthotopic mouse model.

Pancreatic cancer is a lethal disease with few successful treatment options. Recent evidence demonstrates that tumor hypoxia promotes pancreatic tumor invasion, metastasis, and therapy resistance. However, little is known about the complex relationship between hypoxia and the pancreatic tumor microenvironment (TME). In this study, we developed a novel intravital fluorescence microscopy platform with an orthotopic mouse model of pancreatic cancer to study tumor cell hypoxia within the TME in vivo, at cellular resolution, over time. Using a fluorescent BxPC3-DsRed tumor cell line with a hypoxia-response element (HRE)/green fluorescent protein (GFP) reporter, we showed that HRE/GFP is a reliable biomarker of pancreatic tumor hypoxia, responding dynamically and reversibly to changing oxygen concentrations within the TME. We also characterized the spatial relationships between tumor hypoxia, microvasculature, and tumor-associated collagen structures using in vivo second harmonic generation microscopy. This quantitative multimodal imaging platform enables the unprecedented study of hypoxia within the pancreatic TME in vivo.

DORA-compliant measures of research quality and impact to assess the performance of researchers in biomedical institutions: Review of published research, international best practice and Delphi survey.

OBJECTIVE: The San Francisco Declaration on Research Assessment (DORA) advocates for assessing biomedical research quality and impact, yet academic organizations continue to employ traditional measures such as Journal Impact Factor. We aimed to identify and prioritize measures for assessing research quality and impact. METHODS: We conducted a review of published and grey literature to identify measures of research quality and impact, which we included in an online survey. We assembled a panel of researchers and research leaders, and conducted a two-round Delphi survey to prioritize measures rated as high (rated 6 or 7 by ≥ 80% of respondents) or moderate (rated 6 or 7 by ≥ 50% of respondents) importance. RESULTS: We identified 50 measures organized in 8 domains: relevance of the research program, challenges to research program, or productivity, team/open science, funding, innovations, publications, other dissemination, and impact. Rating of measures by 44 panelists (60%) in Round One and 24 (55%) in Round Two of a Delphi survey resulted in consensus on the high importance of 5 measures: research advances existing knowledge, research plan is innovative, an independent body of research (or fundamental role) supported by peer-reviewed research funding, research outputs relevant to discipline, and quality of the content of publications. Five measures achieved consensus on moderate importance: challenges to research productivity, potential to improve health or healthcare, team science, collaboration, and recognition by professional societies or academic bodies. There was high congruence between researchers and research leaders across disciplines. CONCLUSIONS: Our work contributes to the field by identifying 10 DORA-compliant measures of research quality and impact, a more comprehensive and explicit set of measures than prior efforts. Research is needed to identify strategies to overcome barriers of use of DORA-compliant measures, and to "de-implement" traditional measures that do not uphold DORA principles yet are still in use.

Assessing the Accuracy of Bioluminescence Image-Guided Stereotactic Body Radiation Therapy of Orthotopic Pancreatic Tumors Using a Small Animal Irradiator.

Stereotactic body radiation therapy (SBRT) has shown promising results in the treatment of pancreatic cancer and other solid tumors. However, wide adoption of SBRT remains limited largely due to uncertainty about the treatment's optimal fractionation schedules to elicit maximal tumor response while limiting the dose to adjacent structures. A small animal irradiator in combination with a clinically relevant oncological animal model could address these questions. Accurate delivery of X rays to animal tumors may be hampered by suboptimal image-guided targeting of the X-ray beam in vivo. Integration of bioluminescence imaging (BLI) into small animal irradiators in addition to standard cone-beam computed tomography (CBCT) imaging improves target identification and high-precision therapy delivery to deep tumors with poor soft tissue contrast, such as pancreatic tumors. Using bioluminescent BxPC3 pancreatic adenocarcinoma human cells grown orthotopically in mice, we examined the performance of a small animal irradiator equipped with both CBCT and BLI in delivering targeted, hypo-fractionated, multi-beam SBRT. Its targeting accuracy was compared with magnetic resonance imaging (MRI)-guided targeting based on co-registration between CBCT and corresponding sequential magnetic resonance scans, which offer greater soft tissue contrast compared with CT alone. Evaluation of our platform's BLI-guided targeting accuracy was performed by quantifying in vivo changes in bioluminescence signal after treatment as well as staining of ex vivo tissues with γH2AX, Ki67, TUNEL, CD31 and CD11b to assess SBRT treatment effects. Using our platform, we found that BLI-guided SBRT enabled more accurate delivery of X rays to the tumor resulting in greater cancer cell DNA damage and proliferation inhibition compared with MRI-guided SBRT. Furthermore, BLI-guided SBRT allowed higher animal throughput and was more cost effective to use in the preclinical setting than MRI-guided SBRT. Taken together, our preclinical platform could be employed in translational research of SBRT of pancreatic cancer.

Intraoperative fluorescence imaging with aminolevulinic acid detects grossly occult breast cancer: a phase II randomized controlled trial.

BACKGROUND: Re-excision due to positive margins following breast-conserving surgery (BCS) negatively affects patient outcomes and healthcare costs. The inability to visualize margin involvement is a significant challenge in BCS. 5-Aminolevulinic acid hydrochloride (5-ALA HCl), a non-fluorescent oral prodrug, causes intracellular accumulation of fluorescent porphyrins in cancer cells. This single-center Phase II randomized controlled trial evaluated the safety, feasibility, and diagnostic accuracy of a prototype handheld fluorescence imaging device plus 5-ALA for intraoperative visualization of invasive breast carcinomas during BCS. METHODS: Fifty-four patients were enrolled and randomized to receive no 5-ALA or oral 5-ALA HCl (15 or 30 mg/kg). Forty-five patients (n = 15/group) were included in the analysis. Fluorescence imaging of the excised surgical specimen was performed, and biopsies were collected from within and outside the clinically demarcated tumor border of the gross specimen for blinded histopathology. RESULTS: In the absence of 5-ALA, tissue autofluorescence imaging lacked tumor-specific fluorescent contrast. Both 5-ALA doses caused bright red tumor fluorescence, with improved visualization of tumor contrasted against normal tissue autofluorescence. In the 15 mg/kg 5-ALA group, the positive predictive value (PPV) for detecting breast cancer inside and outside the grossly demarcated tumor border was 100.0% and 55.6%, respectively. In the 30 mg/kg 5-ALA group, the PPV was 100.0% and 50.0% inside and outside the demarcated tumor border, respectively. No adverse events were observed, and clinical feasibility of this imaging device-5-ALA combination approach was confirmed. CONCLUSIONS: This is the first known clinical report of visualization of 5-ALA-induced fluorescence in invasive breast carcinoma using a real-time handheld intraoperative fluorescence imaging device. TRIAL REGISTRATION: Clinicaltrials.gov identifier NCT01837225 . Registered 23 April 2013.

New insights into spatio-temporal dynamics of mesenchymal progenitor cell ingress during peri-implant wound healing: Provided by intravital imaging.

Surface topography drives the success of orthopedic and dental implants placed in bone, by directing the biology occurring at the tissue-implant interface. Over the last few decades, striking advancements have been made in the development of novel implant surfaces that enhance bone anchorage to their surfaces through contact osteogenesis: the combination of the two phenomena of recruitment and migration of mesenchymal progenitor cells to the implant surface, and their differentiation into bone-forming cells. While the latter is generally understood, the mechanisms and dynamics underlying the migration and recruitment of such progenitor cells into the wound site have garnered little attention. To address this deficit, we surgically inserted metallic implants with two different surface topographies into the skulls of mice, and then employed real-time spatiotemporal microscopic monitoring of the peri-implant tissue healing to track the ingress of cells. Our results show that nano-topographically complex, in comparison to relatively smooth, implant surfaces profoundly affect recruitment of both endothelial cells, which are essential for angiogenesis, and the mesenchymal progenitor cells that give rise to the reparative tissue stroma. The latter appear concomitantly in the wound site with endothelial cells, from the vascularized areas of the periosteum, and demonstrate a proliferative "bloom" that diminishes with time, although some of these cells differentiate into important stromal cells, pericytes and osteocytes, of the reparative wound. In separate experiments we show, using trajectory plots, that the directionality of migration for both endothelial and perivascular cells can be explained by implant surface dependent release of local cytokine gradients from platelets that would become activated on the implant surfaces during initial blood contact. These findings provide new biological insights into the earliest stages of wound healing, and have broad implications in the application of putative nano-topographically complex biomaterials in many tissue types.

Engineering functional microvessels in synthetic polyurethane random-pore scaffolds by harnessing perfusion flow.

Recently reported biomaterial-based approaches toward prevascularizing tissue constructs rely on biologically or structurally complex scaffolds that are complicated to manufacture and sterilize, and challenging to customize for clinical applications. In the current work, a prevascularization method for soft tissue engineering that uses a non-patterned and non-biological scaffold is proposed. Human fibroblasts and HUVECs were seeded on an ionomeric polyurethane-based hydrogel and cultured for 14 days under medium perfusion. A flow rate of 0.05 mL/min resulted in a greater lumen density in the constructs relative to 0.005 and 0.5 mL/min, indicating the critical importance of flow magnitude in establishing microvessels. Constructs generated at 0.05 mL/min perfusion flow were implanted in a mouse subcutaneous model and intravital imaging was used to characterize host blood perfusion through the construct after 2 weeks. Engineered microvessels were functional (i.e. perfused with host blood and non-leaky) and neovascularization of the construct by host vessels was enhanced relative to non-prevascularized constructs. We report on the first strategy toward engineering functional microvessels in a tissue construct using non-bioactive, non-patterned synthetic polyurethane materials.

In vitro detection of porphyrin-producing wound bacteria with real-time fluorescence imaging.

Aim: Fluorescence imaging can visualize polymicrobial populations in chronic and acute wounds based on porphyrin fluorescence. We investigated the fluorescent properties of specific wound pathogens and the fluorescence detected from bacteria in biofilm. Methods: Utilizing Remel Porphyrin Test Agar, 32 bacterial and four yeast species were examined for red fluorescence under 405 nm violet light illumination. Polymicrobial biofilms, supplemented with δ-aminolevulinic acid, were investigated similarly. Results: A total of 28/32 bacteria, 1/4 yeast species and polymicrobial biofilms produced red fluorescence, in agreement with their known porphyrin production abilities. Conclusion: These results identify common wound pathogens capable of producing porphyrin-specific fluorescence and support clinical observations using fluorescence imaging to detect pathogenic bacteria in chronic wounds.

Intravital Imaging for Tracking of Angiogenesis and Cellular Events Around Surgical Bone Implants.

IMPACT STATEMENT: These new experimental methods allow us to image, and quantify, angiogenesis and perivascular cell dynamics in the endosseous healing compartment. As such, the method is capable of providing a new perspective on, and unique information regarding, healing that occurs around orthopedic and dental implants.

Nanosurfaces modulate the mechanism of peri-implant endosseous healing by regulating neovascular morphogenesis.

Nanosurfaces have improved clinical osseointegration by increasing bone/implant contact. Neovascularization is considered an essential prerequisite to osteogenesis, but no previous reports to our knowledge have examined the effect of surface topography on the spatio-temporal pattern of neovascularization during peri-implant healing. We have developed a cranial window model to study peri-implant healing intravitally over clinically relevant time scales as a function of implant topography. Quantitative intravital confocal imaging reveals that changing the topography (but not chemical composition) of an implant profoundly affects the pattern of peri-implant neovascularization. New vessels develop proximal to the implant and the vascular network matures sooner in the presence of an implant nanosurface. Accelerated angiogenesis can lead to earlier osseointegration through the delivery of osteogenic precursors to, and direct formation of bone on, the implant surface. This study highlights a critical aspect of peri-implant healing, but also informs the biological rationale for the surface design of putative endosseous implant materials.

Improved detection of clinically relevant wound bacteria using autofluorescence image-guided sampling in diabetic foot ulcers.

Clinical wound assessment involves microbiological swabbing of wounds to identify and quantify bacterial species, and to determine microbial susceptibility to antibiotics. The Levine swabbing technique may be suboptimal because it samples only the wound bed, missing other diagnostically relevant areas of the wound, which may contain clinically significant bacteria. Thus, there is a clinical need to improve the reliability of microbiological wound sampling. To address this, a handheld portable autofluorescence (AF) imaging device that detects bacteria in real time, without contrast agents, was developed. Here, we report the results of a clinical study evaluating the use of real-time AF imaging to visualise bacteria in and around the wound bed and to guide swabbing during the clinical assessment of diabetic foot ulcers, compared with the Levine technique. We investigated 33 diabetic foot ulcers (n = 31 patients) and found that AF imaging more accurately identified the presence of moderate and/or heavy bacterial load compared with the Levine technique (accuracy 78% versus 52%, P = 0·048; adjusted diagnostic odds ratio 7·67, P < 0·00022 versus 3·07, P = 0·066) and maximised the effectiveness of bacterial load sampling, with no significant impact on clinical workflow. AF imaging may help clinicians better identify the wound areas with clinically significant bacteria, and maximise sampling of treatment-relevant pathogens.

In Vivo Imaging Reveals Significant Tumor Vascular Dysfunction and Increased Tumor Hypoxia-Inducible Factor-1α Expression Induced by High Single-Dose Irradiation in a Pancreatic Tumor Model.

PURPOSE: To investigate the effect of high-dose irradiation on pancreatic tumor vasculature and microenvironment using in vivo imaging techniques. METHODS AND MATERIALS: A BxPC3 pancreatic tumor xenograft was established in a dorsal skinfold window chamber model and a subcutaneous hind leg model. Tumors were irradiated with a single dose of 4, 12, or 24 Gy. The dorsal skinfold window chamber model was used to assess tumor response, vascular function and permeability, platelet and leukocyte adhesion to the vascular endothelium, and tumor hypoxia for up to 14 days after 24-Gy irradiation. The hind leg model was used to monitor tumor size, hypoxia, and vascularity for up to 65 days after 24-Gy irradiation. Tumors were assessed histologically to validate in vivo observations. RESULTS: In vivo fluorescence imaging revealed temporary vascular dysfunction in tumors irradiated with a single dose of 4 to 24 Gy, but most significantly with a single dose of 24 Gy. Vascular functional recovery was observed by 14 days after irradiation in a dose-dependent manner. Furthermore, irradiation with 24 Gy caused platelet and leukocyte adhesion to the vascular endothelium within hours to days after irradiation. Vascular permeability was significantly higher in irradiated tumors compared with nonirradiated controls 14 days after irradiation. This observation corresponded with increased expression of hypoxia-inducible factor-1α in irradiated tumors. In the hind leg model, irradiation with a single dose of 24 Gy led to tumor growth delay, followed by tumor regrowth. CONCLUSIONS: Irradiation of the BxPC3 tumors with a single dose of 24 Gy caused transient vascular dysfunction and increased expression of hypoxia-inducible factor-1α. Such biological changes may impact tumor response to high single-dose and hypofractionated irradiation, and further investigations are needed to better understand the clinical outcomes of stereotactic body radiation therapy.

Hybrid Manganese Dioxide Nanoparticles Potentiate Radiation Therapy by Modulating Tumor Hypoxia.

Hypoxia in the tumor microenvironment (TME) mediates resistance to radiotherapy and contributes to poor prognosis in patients receiving radiotherapy. Here we report the design of clinically suitable formulations of hybrid manganese dioxide (MnO2) nanoparticles (MDNP) using biocompatible materials to reoxygenate the TME by reacting with endogenous H2O2 MDNP containing hydrophilic terpolymer-protein-MnO2 or hydrophobic polymer-lipid-MnO2 provided different oxygen generation rates in the TME relevant to different clinical settings. In highly hypoxic murine or human xenograft breast tumor models, we found that administering either MDNP formulation before radiotherapy modulated tumor hypoxia and increased radiotherapy efficacy, acting to reduce tumor growth, VEGF expression, and vascular density. MDNP treatment also increased apoptosis and DNA double strand breaks, increasing median host survival 3- to 5-fold. Notably, in the murine model, approximately 40% of tumor-bearing mice were tumor-free after a single treatment with MDNPs plus radiotherapy at a 2.5-fold lower dose than required to achieve the same curative treatment without MDNPs. Overall, our findings offer a preclinical proof of concept for the use of MDNP formulations as effective radiotherapy adjuvants. Cancer Res; 76(22); 6643-56. ©2016 AACR.

The Necrosis-Avid Small Molecule HQ4-DTPA as a Multimodal Imaging Agent for Monitoring Radiation Therapy-Induced Tumor Cell Death.

PURPOSE: Most effective antitumor therapies induce tumor cell death. Non-invasive, rapid and accurate quantitative imaging of cell death is essential for monitoring early response to antitumor therapies. To facilitate this, we previously developed a biocompatible necrosis-avid near-infrared fluorescence (NIRF) imaging probe, HQ4, which was radiolabeled with 111Indium-chloride (111In-Cl3) via the chelate diethylene triamine pentaacetic acid (DTPA), to enable clinical translation. The aim of the present study was to evaluate the application of HQ4-DTPA for monitoring tumor cell death induced by radiation therapy. Apart from its NIRF and radioactive properties, HQ4-DTPA was also tested as a photoacoustic imaging probe to evaluate its performance as a multimodal contrast agent for superficial and deep tissue imaging. MATERIALS AND METHODS: Radiation-induced tumor cell death was examined in a xenograft mouse model of human breast cancer (MCF-7). Tumors were irradiated with three fractions of 9 Gy each. HQ4-DTPA was injected intravenously after the last irradiation, NIRF and photoacoustic imaging of the tumors were performed at 12, 20, and 40 h after injection. Changes in probe accumulation in the tumors were measured in vivo, and ex vivo histological analysis of excised tumors was performed at experimental endpoints. In addition, biodistribution of radiolabeled [111In]DTPA-HQ4 was assessed using hybrid single-photon emission computed tomography-computed tomography (SPECT-CT) at the same time points. RESULTS: In vivo NIRF imaging demonstrated a significant difference in probe accumulation between control and irradiated tumors at all time points after injection. A similar trend was observed using in vivo photoacoustic imaging, which was validated by ex vivo tissue fluorescence and photoacoustic imaging. Serial quantitative radioactivity measurements of probe biodistribution further demonstrated increased probe accumulation in irradiated tumors. CONCLUSION: HQ4-DTPA has high specificity for dead cells in vivo, potentiating its use as a contrast agent for determining the relative level of tumor cell death following radiation therapy using NIRF, photoacoustic imaging and SPECT in vivo. Initial preclinical results are promising and indicate the need for further evaluation in larger cohorts. If successful, such studies may help develop a new multimodal method for non-invasive and dynamic deep tissue imaging of treatment-induced cell death to quantitatively assess therapeutic response in patients.

Can Imaging Put the "Advanced" Back in Advanced Wound Care?

An effective, scientifically validated, diagnostic tool helps clinicians make better, timely, and more objective medical decisions in the care of their patients. Today, the need for such tools is especially urgent in the field of wound care where patient-centric care is the goal, under ever tightening clinical budget constraints. In an era of countless "innovative" treatment options, that is, advanced dressings, negative pressure devices, and various debridement instruments available to the wound care clinical team, one area that has arguably languished in the past decade has been innovation in wound diagnostics. Whereas medical imaging is a mainstay in the diagnostic toolkit across many other medical fields (oncology, neurology, gastroenterology, orthopedics, etc.), the field of wound care has yet to realize the full potential that advances in imaging technologies have to offer the clinician. In this issue, the first of a series in wound imaging and diagnostics, four articles have been assembled, highlighting some of the recent advances in wound imaging technologies.

Pre-clinical Evaluation of a Cyanine-Based SPECT Probe for Multimodal Tumor Necrosis Imaging.

PURPOSE: Recently we showed that a number of carboxylated near-infrared fluorescent (NIRF) cyanine dyes possess strong necrosis avid properties in vitro as well as in different mouse models of spontaneous and therapy-induced tumor necrosis, indicating their potential use for cancer diagnostic- and prognostic purposes. In the previous study, the detection of the cyanines was achieved by whole body optical imaging, a technique that, due to the limited penetration of near-infrared light, is not suitable for investigations deeper than 1 cm within the human body. Therefore, in order to facilitate clinical translation, the purpose of the present study was to generate a necrosis avid cyanine-based NIRF probe that could also be used for single photon emission computed tomography (SPECT). For this, the necrosis avid NIRF cyanine HQ4 was radiolabeled with 111indium, via the chelate diethylene triamine pentaacetic acid (DTPA). PROCEDURES: The necrosis avid properties of the radiotracer [111In]DTPA-HQ4 were examined in vitro and in vivo in different breast tumor models in mice using SPECT and optical imaging. Moreover, biodistribution studies were performed to examine the pharmacokinetics of the probe in vivo. RESULTS: Using optical imaging and radioactivity measurements, in vitro, we showed selective accumulation of [111In]DTPA-HQ4 in dead cells. Using SPECT and in biodistribution studies, the necrosis avidity of the radiotracer was confirmed in a 4T1 mouse breast cancer model of spontaneous tumor necrosis and in a MCF-7 human breast cancer model of chemotherapy-induced tumor necrosis. CONCLUSIONS: The radiotracer [111In]DTPA-HQ4 possessed strong and selective necrosis avidity in vitro and in various mouse models of tumor necrosis in vivo, indicating its potential to be clinically applied for diagnostic purposes and to monitor anti-cancer treatment efficacy.

Handheld fluorescence imaging device detects subclinical wound infection in an asymptomatic patient with chronic diabetic foot ulcer: a case report.

Chronic wounds are a significant burden to global patient and health care infrastructures, and there is a need for better methods of early wound diagnosis and treatment. Traditional diagnosis of chronic wound infection by pathogenic bacteria, using clinical signs and symptoms, is based on visual inspection under white light and microbiological sampling (e.g. swabbing and/or biopsy) of the wound, which are subjective and suboptimal. Diagnosing microbial infection based on traditional clinical signs and symptoms in wounds of asymptomatic patients is especially challenging at the bedside. Bacteria are invisible to the unaided eye and wound sampling for diagnostic testing can cause unacceptable delays in diagnosis and treatment. To address this problem, we developed a new prototype handheld, portable fluorescence imaging device that enables non-contact, real-time, high-resolution visualisation of pathogenic bacteria and tissues in wounds. Herein, we report the clinical use of this imaging device in detecting subsurface heavy bacterial load and subclinical local infection in an asymptomatic 50-year-old patient with a non-healing diabetic foot ulcer.

Emerging Applications for Optically Enabled Intravital Microscopic Imaging in Radiobiology.

Radiation therapy is an effective cancer treatment used in over 50% of cancer patients. Preclinical research in radiobiology plays a major role in influencing the translation of radiotherapy-based treatment strategies into clinical practice. Studies have demonstrated that various components of tumors and their microenvironments, including vasculature, immune and stem cells, and stromal cells, can influence the response of solid tumors to radiation. Optically enabled imaging techniques used in experimental animal models of cancer offer a unique and powerful way to quantitatively track spatiotemporal changes in these tumor components in vivo at macro-, meso-, and microscopic resolutions following radiotherapy. In this review, we discuss the role of both well-established and emerging intravital microscopy techniques for studying tumors and their microenvironment in vivo, in response to irradiation. The development and application of new animal models, small animal microirradiation technologies, and multimodal optically enabled intravital microscopy techniques are emphasized within the framework of preclinical radiobiology research. We also comment on the potential influence that these newer imaging techniques may have on the clinical translation of new preclinical radiobiology discoveries.

Optically-tracked handheld fluorescence imaging platform for monitoring skin response in the management of soft tissue sarcoma.

Standard clinical management of extremity soft tissue sarcomas includes surgery with radiation therapy. Wound complications (WCs) arising from treatment may occur due to bacterial infection and tissue breakdown. The ability to detect changes in these parameters during treatment may lead to earlier interventions that mitigate WCs. We describe the use of a new system composed of an autofluorescence imaging device and an optical three-dimensional tracking system to detect and coregister the presence of bacteria with radiation doses. The imaging device visualized erythema using white light and detected bacterial autofluorescence using 405-nm excitation light. Its position was tracked relative to the patient using IR reflective spheres and registration to the computed tomography coordinates. Image coregistration software was developed to spatially overlay radiation treatment plans and dose distributions on the white light and autofluorescence images of the surgical site. We describe the technology, its use in the operating room, and standard operating procedures, as well as demonstrate technical feasibility and safety intraoperatively. This new clinical tool may help identify patients at greater risk of developing WCs and investigate correlations between radiation dose, skin response, and changes in bacterial load as biomarkers associated with WCs.