Computer assisted analysis of contrast enhanced ultrasound images for quantification in vascular diseases

Contrast enhanced ultrasound (CEUS) with microbubble contrast agents has shown great potential in imaging microvasculature, quantifying perfusion and hence detecting vascular diseases. However, most existing perfusion quantification methods based on image intensity, and are susceptible to confounding factors such as attenuation artefacts. Improving reproducibility is also a key challenge to clinical translation. Therefore, this thesis aims at developing attenuation correction and quantification techniques in CEUS with applications for detection and quantification of microvascular flow / perfusion. Firstly, a technique for automatic correction of attenuation effects in vascular imaging was developed and validated on a tissue mimicking phantom. The application of this technique to studying contrast enhancement of carotid adventitial vasa vasorum as a biomarker of radiation-induced atherosclerosis was demonstrated. The results showed great potential in reducing attenuation artefact and improve quantification in CEUS of carotid arteries. Furthermore, contrast intensity was shown to significantly increase in irradiated carotid arteries and could be a useful imaging biomarker for radiation-induced atherosclerosis. Secondly, a robust and automated tool for quantification of microbubble identification in CEUS image sequences using a temporal and spatial analysis was developed and validated on a flow phantom. The application of this technique to evaluate human musculoskeletal microcirculation with contrast enhanced ultrasound was demonstrated. The results showed an excellent accuracy and repeatability in quantifying active vascular density. It has great potential for clinical translation in the assessment of lower limb perfusion. Finally, a new bubble activity identification and quantification technique based on differential intensity projection in CEUS was developed and demonstrated with an in-vivo study, and applied to the quantification of intraplaque neovascularisation in an irradiated carotid artery of patients who were previously treated for head and neck cancer. The results showed a significantly more specific identification of bubble signals and had good agreement between the differential intensity-based technique and clinical visual assessment. This technique has potential to assist clinicians to diagnose and monitor intraplque neovascularisation.Open Acces

Automatic Spatiotemporal Analysis of Cardiac Image Series

RÉSUMÉ À ce jour, les maladies cardiovasculaires demeurent au premier rang des principales causes de décès en Amérique du Nord. Chez l’adulte et au sein de populations de plus en plus jeunes, la soi-disant épidémie d’obésité entraînée par certaines habitudes de vie tels que la mauvaise alimentation, le manque d’exercice et le tabagisme est lourde de conséquences pour les personnes affectées, mais aussi sur le système de santé. La principale cause de morbidité et de mortalité chez ces patients est l’athérosclérose, une accumulation de plaque à l’intérieur des vaisseaux sanguins à hautes pressions telles que les artères coronaires. Les lésions athérosclérotiques peuvent entraîner l’ischémie en bloquant la circulation sanguine et/ou en provoquant une thrombose. Cela mène souvent à de graves conséquences telles qu’un infarctus. Outre les problèmes liés à la sténose, les parois artérielles des régions criblées de plaque augmentent la rigidité des parois vasculaires, ce qui peut aggraver la condition du patient. Dans la population pédiatrique, la pathologie cardiovasculaire acquise la plus fréquente est la maladie de Kawasaki. Il s’agit d’une vasculite aigüe pouvant affecter l’intégrité structurale des parois des artères coronaires et mener à la formation d’anévrismes. Dans certains cas, ceux-ci entravent l’hémodynamie artérielle en engendrant une perfusion myocardique insuffisante et en activant la formation de thromboses. Le diagnostic de ces deux maladies coronariennes sont traditionnellement effectués à l’aide d’angiographies par fluoroscopie. Pendant ces examens paracliniques, plusieurs centaines de projections radiographiques sont acquises en séries suite à l’infusion artérielle d’un agent de contraste. Ces images révèlent la lumière des vaisseaux sanguins et la présence de lésions potentiellement pathologiques, s’il y a lieu. Parce que les séries acquises contiennent de l’information très dynamique en termes de mouvement du patient volontaire et involontaire (ex. battements cardiaques, respiration et déplacement d’organes), le clinicien base généralement son interprétation sur une seule image angiographique où des mesures géométriques sont effectuées manuellement ou semi-automatiquement par un technicien en radiologie. Bien que l’angiographie par fluoroscopie soit fréquemment utilisé partout dans le monde et souvent considéré comme l’outil de diagnostic “gold-standard” pour de nombreuses maladies vasculaires, la nature bidimensionnelle de cette modalité d’imagerie est malheureusement très limitante en termes de spécification géométrique des différentes régions pathologiques. En effet, la structure tridimensionnelle des sténoses et des anévrismes ne peut pas être pleinement appréciée en 2D car les caractéristiques observées varient selon la configuration angulaire de l’imageur. De plus, la présence de lésions affectant les artères coronaires peut ne pas refléter la véritable santé du myocarde, car des mécanismes compensatoires naturels (ex. vaisseaux----------ABSTRACT Cardiovascular disease continues to be the leading cause of death in North America. In adult and, alarmingly, ever younger populations, the so-called obesity epidemic largely driven by lifestyle factors that include poor diet, lack of exercise and smoking, incurs enormous stresses on the healthcare system. The primary cause of serious morbidity and mortality for these patients is atherosclerosis, the build up of plaque inside high pressure vessels like the coronary arteries. These lesions can lead to ischemic disease and may progress to precarious blood flow blockage or thrombosis, often with infarction or other severe consequences. Besides the stenosis-related outcomes, the arterial walls of plaque-ridden regions manifest increased stiffness, which may exacerbate negative patient prognosis. In pediatric populations, the most prevalent acquired cardiovascular pathology is Kawasaki disease. This acute vasculitis may affect the structural integrity of coronary artery walls and progress to aneurysmal lesions. These can hinder the blood flow’s hemodynamics, leading to inadequate downstream perfusion, and may activate thrombus formation which may lead to precarious prognosis. Diagnosing these two prominent coronary artery diseases is traditionally performed using fluoroscopic angiography. Several hundred serial x-ray projections are acquired during selective arterial infusion of a radiodense contrast agent, which reveals the vessels’ luminal area and possible pathological lesions. The acquired series contain highly dynamic information on voluntary and involuntary patient movement: respiration, organ displacement and heartbeat, for example. Current clinical analysis is largely limited to a single angiographic image where geometrical measures will be performed manually or semi-automatically by a radiological technician. Although widely used around the world and generally considered the gold-standard diagnosis tool for many vascular diseases, the two-dimensional nature of this imaging modality is limiting in terms of specifying the geometry of various pathological regions. Indeed, the 3D structures of stenotic or aneurysmal lesions may not be fully appreciated in 2D because their observable features are dependent on the angular configuration of the imaging gantry. Furthermore, the presence of lesions in the coronary arteries may not reflect the true health of the myocardium, as natural compensatory mechanisms may obviate the need for further intervention. In light of this, cardiac magnetic resonance perfusion imaging is increasingly gaining attention and clinical implementation, as it offers a direct assessment of myocardial tissue viability following infarction or suspected coronary artery disease. This type of modality is plagued, however, by motion similar to that present in fluoroscopic imaging. This issue predisposes clinicians to laborious manual intervention in order to align anatomical structures in sequential perfusion frames, thus hindering automation o

Analysis of contrast-enhanced medical images.

Early detection of human organ diseases is of great importance for the accurate diagnosis and institution of appropriate therapies. This can potentially prevent progression to end-stage disease by detecting precursors that evaluate organ functionality. In addition, it also assists the clinicians for therapy evaluation, tracking diseases progression, and surgery operations. Advances in functional and contrast-enhanced (CE) medical images enabled accurate noninvasive evaluation of organ functionality due to their ability to provide superior anatomical and functional information about the tissue-of-interest. The main objective of this dissertation is to develop a computer-aided diagnostic (CAD) system for analyzing complex data from CE magnetic resonance imaging (MRI). The developed CAD system has been tested in three case studies: (i) early detection of acute renal transplant rejection, (ii) evaluation of myocardial perfusion in patients with ischemic heart disease after heart attack; and (iii), early detection of prostate cancer. However, developing a noninvasive CAD system for the analysis of CE medical images is subject to multiple challenges, including, but are not limited to, image noise and inhomogeneity, nonlinear signal intensity changes of the images over the time course of data acquisition, appearances and shape changes (deformations) of the organ-of-interest during data acquisition, determination of the best features (indexes) that describe the perfusion of a contrast agent (CA) into the tissue. To address these challenges, this dissertation focuses on building new mathematical models and learning techniques that facilitate accurate analysis of CAs perfusion in living organs and include: (i) accurate mathematical models for the segmentation of the object-of-interest, which integrate object shape and appearance features in terms of pixel/voxel-wise image intensities and their spatial interactions; (ii) motion correction techniques that combine both global and local models, which exploit geometric features, rather than image intensities to avoid problems associated with nonlinear intensity variations of the CE images; (iii) fusion of multiple features using the genetic algorithm. The proposed techniques have been integrated into CAD systems that have been tested in, but not limited to, three clinical studies. First, a noninvasive CAD system is proposed for the early and accurate diagnosis of acute renal transplant rejection using dynamic contrast-enhanced MRI (DCE-MRI). Acute rejection–the immunological response of the human immune system to a foreign kidney–is the most sever cause of renal dysfunction among other diagnostic possibilities, including acute tubular necrosis and immune drug toxicity. In the U.S., approximately 17,736 renal transplants are performed annually, and given the limited number of donors, transplanted kidney salvage is an important medical concern. Thus far, biopsy remains the gold standard for the assessment of renal transplant dysfunction, but only as the last resort because of its invasive nature, high cost, and potential morbidity rates. The diagnostic results of the proposed CAD system, based on the analysis of 50 independent in-vivo cases were 96% with a 95% confidence interval. These results clearly demonstrate the promise of the proposed image-based diagnostic CAD system as a supplement to the current technologies, such as nuclear imaging and ultrasonography, to determine the type of kidney dysfunction. Second, a comprehensive CAD system is developed for the characterization of myocardial perfusion and clinical status in heart failure and novel myoregeneration therapy using cardiac first-pass MRI (FP-MRI). Heart failure is considered the most important cause of morbidity and mortality in cardiovascular disease, which affects approximately 6 million U.S. patients annually. Ischemic heart disease is considered the most common underlying cause of heart failure. Therefore, the detection of the heart failure in its earliest forms is essential to prevent its relentless progression to premature death. While current medical studies focus on detecting pathological tissue and assessing contractile function of the diseased heart, this dissertation address the key issue of the effects of the myoregeneration therapy on the associated blood nutrient supply. Quantitative and qualitative assessment in a cohort of 24 perfusion data sets demonstrated the ability of the proposed framework to reveal regional perfusion improvements with therapy, and transmural perfusion differences across the myocardial wall; thus, it can aid in follow-up on treatment for patients undergoing the myoregeneration therapy. Finally, an image-based CAD system for early detection of prostate cancer using DCE-MRI is introduced. Prostate cancer is the most frequently diagnosed malignancy among men and remains the second leading cause of cancer-related death in the USA with more than 238,000 new cases and a mortality rate of about 30,000 in 2013. Therefore, early diagnosis of prostate cancer can improve the effectiveness of treatment and increase the patient’s chance of survival. Currently, needle biopsy is the gold standard for the diagnosis of prostate cancer. However, it is an invasive procedure with high costs and potential morbidity rates. Additionally, it has a higher possibility of producing false positive diagnosis due to relatively small needle biopsy samples. Application of the proposed CAD yield promising results in a cohort of 30 patients that would, in the near future, represent a supplement of the current technologies to determine prostate cancer type. The developed techniques have been compared to the state-of-the-art methods and demonstrated higher accuracy as shown in this dissertation. The proposed models (higher-order spatial interaction models, shape models, motion correction models, and perfusion analysis models) can be used in many of today’s CAD applications for early detection of a variety of diseases and medical conditions, and are expected to notably amplify the accuracy of CAD decisions based on the automated analysis of CE images

Characterization of carotid artery plaques using noninvasive vascular ultrasound elastography

L'athérosclérose est une maladie vasculaire complexe qui affecte la paroi des artères (par l'épaississement) et les lumières (par la formation de plaques). La rupture d'une plaque de l'artère carotide peut également provoquer un accident vasculaire cérébral ischémique et des complications. Bien que plusieurs modalités d'imagerie médicale soient actuellement utilisées pour évaluer la stabilité d'une plaque, elles présentent des limitations telles que l'irradiation, les propriétés invasives, une faible disponibilité clinique et un coût élevé. L'échographie est une méthode d'imagerie sûre qui permet une analyse en temps réel pour l'évaluation des tissus biologiques. Il est intéressant et prometteur d’appliquer une échographie vasculaire pour le dépistage et le diagnostic précoces des plaques d’artère carotide. Cependant, les ultrasons vasculaires actuels identifient uniquement la morphologie d'une plaque en termes de luminosité d'écho ou l’impact de cette plaque sur les caractéristiques de l’écoulement sanguin, ce qui peut ne pas être suffisant pour diagnostiquer l’importance de la plaque. La technique d’élastographie vasculaire non-intrusive (« noninvasive vascular elastography (NIVE) ») a montré le potentiel de détermination de la stabilité d'une plaque. NIVE peut déterminer le champ de déformation de la paroi vasculaire en mouvement d’une artère carotide provoqué par la pulsation cardiaque naturelle. En raison des différences de module de Young entre les différents tissus des vaisseaux, différents composants d’une plaque devraient présenter différentes déformations, caractérisant ainsi la stabilité de la plaque. Actuellement, les performances et l’efficacité numérique sous-optimales limitent l’acceptation clinique de NIVE en tant que méthode rapide et efficace pour le diagnostic précoce des plaques vulnérables. Par conséquent, il est nécessaire de développer NIVE en tant qu’outil d’imagerie non invasif, rapide et économique afin de mieux caractériser la vulnérabilité liée à la plaque. La procédure à suivre pour effectuer l’analyse NIVE consiste en des étapes de formation et de post-traitement d’images. Cette thèse vise à améliorer systématiquement la précision de ces deux aspects de NIVE afin de faciliter la prédiction de la vulnérabilité de la plaque carotidienne. Le premier effort de cette thèse a été dédié à la formation d'images (Chapitre 5). L'imagerie par oscillations transversales a été introduite dans NIVE. Les performances de l’imagerie par oscillations transversales couplées à deux estimateurs de contrainte fondés sur un modèle de déformation fine, soit l’ « affine phase-based estimator (APBE) » et le « Lagrangian speckle model estimator (LSME) », ont été évaluées. Pour toutes les études de simulation et in vitro de ce travail, le LSME sans imagerie par oscillation transversale a surperformé par rapport à l'APBE avec imagerie par oscillations transversales. Néanmoins, des estimations de contrainte principales comparables ou meilleures pourraient être obtenues avec le LSME en utilisant une imagerie par oscillations transversales dans le cas de structures tissulaires complexes et hétérogènes. Lors de l'acquisition de signaux ultrasonores pour la formation d'images, des mouvements hors du plan perpendiculaire au plan de balayage bidimensionnel (2-D) existent. Le deuxième objectif de cette thèse était d'évaluer l'influence des mouvements hors plan sur les performances du NIVE 2-D (Chapitre 6). À cette fin, nous avons conçu un dispositif expérimental in vitro permettant de simuler des mouvements hors plan de 1 mm, 2 mm et 3 mm. Les résultats in vitro ont montré plus d'artefacts d'estimation de contrainte pour le LSME avec des amplitudes croissantes de mouvements hors du plan principal de l’image. Malgré tout, nous avons néanmoins obtenu des estimations de déformations robustes avec un mouvement hors plan de 2.0 mm (coefficients de corrélation supérieurs à 0.85). Pour un jeu de données cliniques de 18 participants présentant une sténose de l'artère carotide, nous avons proposé d'utiliser deux jeux de données d'analyses sur la même plaque carotidienne, soit des images transversales et longitudinales, afin de déduire les mouvements hors plan (qui se sont avérés de 0.25 mm à 1.04 mm). Les résultats cliniques ont montré que les estimations de déformations restaient reproductibles pour toutes les amplitudes de mouvement, puisque les coefficients de corrélation inter-images étaient supérieurs à 0.70 et que les corrélations croisées normalisées entre les images radiofréquences étaient supérieures à 0.93, ce qui a permis de démontrer une plus grande confiance lors de l'analyse de jeu de données cliniques de plaques carotides à l'aide du LSME. Enfin, en ce qui concerne le post-traitement des images, les algorithmes NIVE doivent estimer les déformations des parois des vaisseaux à partir d’images reconstituées dans le but d’identifier les tissus mous et durs. Ainsi, le dernier objectif de cette thèse était de développer un algorithme d'estimation de contrainte avec une résolution de la taille d’un pixel ainsi qu'une efficacité de calcul élevée pour l'amélioration de la précision de NIVE (Chapitre 7). Nous avons proposé un estimateur de déformation de modèle fragmenté (SMSE) avec lequel le champ de déformation dense est paramétré avec des descriptions de transformées en cosinus discret, générant ainsi des composantes de déformations affines (déformations axiales et latérales et en cisaillement) sans opération mathématique de dérivées. En comparant avec le LSME, le SMSE a réduit les erreurs d'estimation lors des tests de simulations, ainsi que pour les mesures in vitro et in vivo. De plus, la faible mise en oeuvre de la méthode SMSE réduit de 4 à 25 fois le temps de traitement par rapport à la méthode LSME pour les simulations, les études in vitro et in vivo, ce qui pourrait permettre une implémentation possible de NIVE en temps réel.Atherosclerosis is a complex vascular disease that affects artery walls (by thickening) and lumens (by plaque formation). The rupture of a carotid artery plaque may also induce ischemic stroke and complications. Despite the use of several medical imaging modalities to evaluate the stability of a plaque, they present limitations such as irradiation, invasive property, low clinical availability and high cost. Ultrasound is a safe imaging method with a real time capability for assessment of biological tissues. It is clinically used for early screening and diagnosis of carotid artery plaques. However, current vascular ultrasound technologies only identify the morphology of a plaque in terms of echo brightness or the impact of the vessel narrowing on flow properties, which may not be sufficient for optimum diagnosis. Noninvasive vascular elastography (NIVE) has been shown of interest for determining the stability of a plaque. Specifically, NIVE can determine the strain field of the moving vessel wall of a carotid artery caused by the natural cardiac pulsation. Due to Young’s modulus differences among different vessel tissues, different components of a plaque can be detected as they present different strains thereby potentially helping in characterizing the plaque stability. Currently, sub-optimum performance and computational efficiency limit the clinical acceptance of NIVE as a fast and efficient method for the early diagnosis of vulnerable plaques. Therefore, there is a need to further develop NIVE as a non-invasive, fast and low computational cost imaging tool to better characterize the plaque vulnerability. The procedure to perform NIVE analysis consists in image formation and image post-processing steps. This thesis aimed to systematically improve the accuracy of these two aspects of NIVE to facilitate predicting carotid plaque vulnerability. The first effort of this thesis has been targeted on improving the image formation (Chapter 5). Transverse oscillation beamforming was introduced into NIVE. The performance of transverse oscillation imaging coupled with two model-based strain estimators, the affine phase-based estimator (APBE) and the Lagrangian speckle model estimator (LSME), were evaluated. For all simulations and in vitro studies, the LSME without transverse oscillation imaging outperformed the APBE with transverse oscillation imaging. Nonetheless, comparable or better principal strain estimates could be obtained with the LSME using transverse oscillation imaging in the case of complex and heterogeneous tissue structures. During the acquisition of ultrasound signals for image formation, out-of-plane motions which are perpendicular to the two-dimensional (2-D) scan plane are existing. The second objective of this thesis was to evaluate the influence of out-of-plane motions on the performance of 2-D NIVE (Chapter 6). For this purpose, we designed an in vitro experimental setup to simulate out-of-plane motions of 1 mm, 2 mm and 3 mm. The in vitro results showed more strain estimation artifacts for the LSME with increasing magnitudes of out-of-plane motions. Even so, robust strain estimations were nevertheless obtained with 2.0 mm out-of-plane motion (correlation coefficients higher than 0.85). For a clinical dataset of 18 participants with carotid artery stenosis, we proposed to use two datasets of scans on the same carotid plaque, one cross-sectional and the other in a longitudinal view, to deduce the out-of-plane motions (estimated to be ranging from 0.25 mm to 1.04 mm). Clinical results showed that strain estimations remained reproducible for all motion magnitudes since inter-frame correlation coefficients were higher than 0.70, and normalized cross-correlations between radiofrequency images were above 0.93, which indicated that confident motion estimations can be obtained when analyzing clinical dataset of carotid plaques using the LSME. Finally, regarding the image post-processing component of NIVE algorithms to estimate strains of vessel walls from reconstructed images with the objective of identifying soft and hard tissues, we developed a strain estimation method with a pixel-wise resolution as well as a high computation efficiency for improving NIVE (Chapter 7). We proposed a sparse model strain estimator (SMSE) for which the dense strain field is parameterized with Discrete Cosine Transform descriptions, thereby deriving affine strain components (axial and lateral strains and shears) without mathematical derivative operations. Compared with the LSME, the SMSE reduced estimation errors in simulations, in vitro and in vivo tests. Moreover, the sparse implementation of the SMSE reduced the processing time by a factor of 4 to 25 compared with the LSME based on simulations, in vitro and in vivo results, which is suggesting a possible implementation of NIVE in real time

Preoperative Systems for Computer Aided Diagnosis based on Image Registration: Applications to Breast Cancer and Atherosclerosis

Computer Aided Diagnosis (CAD) systems assist clinicians including radiologists and cardiologists to detect abnormalities and highlight conspicuous possible disease. Implementing a pre-operative CAD system contains a framework that accepts related technical as well as clinical parameters as input by analyzing the predefined method and demonstrates the prospective output. In this work we developed the Computer Aided Diagnostic System for biomedical imaging analysis of two applications on Breast Cancer and Atherosclerosis. The aim of the first CAD application is to optimize the registration strategy specifically for Breast Dynamic Infrared Imaging and to make it user-independent. Base on the fact that automated motion reduction in dynamic infrared imaging is on demand in clinical applications, since movement disarranges time-temperature series of each pixel, thus originating thermal artifacts that might bias the clinical decision. All previously proposed registration methods are feature based algorithms requiring manual intervention. We implemented and evaluated 3 different 3D time-series registration methods: 1. Linear affine, 2. Non-linear Bspline, 3. Demons applied to 12 datasets of healthy breast thermal images. The results are evaluated through normalized mutual information with average values of 0.70±0.03, 0.74±0.03 and 0.81±0.09 (out of 1) for Affine, BSpline and Demons registration, respectively, as well as breast boundary overlap and Jacobian determinant of the deformation field. The statistical analysis of the results showed that symmetric diffeomorphic Demons registration method outperforms also with the best breast alignment and non-negative Jacobian values which guarantee image similarity and anatomical consistency of the transformation, due to homologous forces enforcing the pixel geometric disparities to be shortened on all the frames. We propose Demons registration as an effective technique for time-series dynamic infrared registration, to stabilize the local temperature oscillation. The aim of the second implemented CAD application is to assess contribution of calcification in plaque vulnerability and wall rupture and to find its maximum resistance before break in image-based models of carotid artery stenting. The role of calcification inside fibroatheroma during carotid artery stenting operation is controversial in which cardiologists face two major problems during the placement: (i) “plaque protrusion” (i.e. elastic fibrous caps containing early calcifications that penetrate inside the stent); (ii) “plaque vulnerability” (i.e. stiff plaques with advanced calcifications that break the arterial wall or stent). Finite Element Analysis was used to simulate the balloon and stent expansion as a preoperative patient-specific virtual framework. A nonlinear static structural analysis was performed on 20 patients acquired using in vivo MDCT angiography. The Agatston Calcium score was obtained for each patient and subject-specific local Elastic Modulus (EM) was calculated. The in silico results showed that by imposing average ultimate external load of 1.1MPa and 2.3MPa on balloon and stent respectively, average ultimate stress of 55.7±41.2kPa and 171±41.2kPa are obtained on calcifications. The study reveals that a significant positive correlation (R=0.85, p<0.0001) exists on stent expansion between EM of calcification and ultimate stress as well as Plaque Wall Stress (PWS) (R=0.92, p<0.0001), comparing to Ca score that showed insignificant associations with ultimate stress (R=0.44, p=0.057) and PWS (R=0.38, p=0.103), suggesting minor impact of Ca score in plaque rupture. These average data are in good agreement with results obtained by other research groups and we believe this approach enriches the arsenal of tools available for pre-operative prediction of carotid artery stenting procedure in the presence of calcified plaques

Super-Resolution Contrast Enhanced Ultrasound Methodology for the Identification of in-Vivo Vascular Dynamics in 2D

\u3cp\u3eObjectives The aim of this study was to provide an ultrasound-based super-resolution methodology that can be implemented using clinical 2-dimensional ultrasound equipment and standard contrast-enhanced ultrasound modes. In addition, the aim is to achieve this for true-to-life patient imaging conditions, including realistic examination times of a few minutes and adequate image penetration depths that can be used to scan entire organs without sacrificing current super-resolution ultrasound imaging performance. Methods Standard contrast-enhanced ultrasound was used along with bolus or infusion injections of SonoVue (Bracco, Geneva, Switzerland) microbubble (MB) suspensions. An image analysis methodology, translated from light microscopy algorithms, was developed for use with ultrasound contrast imaging video data. New features that are tailored for ultrasound contrast image data were developed for MB detection and segmentation, so that the algorithm can deal with single and overlapping MBs. The method was tested initially on synthetic data, then with a simple microvessel phantom, and then with in vivo ultrasound contrast video loops from sheep ovaries. Tracks detailing the vascular structure and corresponding velocity map of the sheep ovary were reconstructed. Images acquired from light microscopy, optical projection tomography, and optical coherence tomography were compared with the vasculature network that was revealed in the ultrasound contrast data. The final method was applied to clinical prostate data as a proof of principle. Results Features of the ovary identified in optical modalities mentioned previously were also identified in the ultrasound super-resolution density maps. Follicular areas, follicle wall, vessel diameter, and tissue dimensions were very similar. An approximately 8.5-fold resolution gain was demonstrated in vessel width, as vessels of width down to 60 μm were detected and verified (λ = 514 μm). Best agreement was found between ultrasound measurements and optical coherence tomography with 10% difference in the measured vessel widths, whereas ex vivo microscopy measurements were significantly lower by 43% on average. The results were mostly achieved using video loops of under 2-minute duration that included respiratory motion. A feasibility study on a human prostate showed good agreement between density and velocity ultrasound maps with the histological evaluation of the location of a tumor. Conclusions The feasibility of a 2-dimensional contrast-enhanced ultrasound-based super-resolution method was demonstrated using in vitro, synthetic and in vivo animal data. The method reduces the examination times to a few minutes using state-of-the-art ultrasound equipment and can provide super-resolution maps for an entire prostate with similar resolution to that achieved in other studies.\u3c/p\u3