The focus of this thesis is the improvement of acquisition and processing of Mueller matrix polarimetry using a confocal scanning laser ophthalmoscope (CSLO) and the application of Mueller matrix polarimetry to image the retina. Stepper motors were incorporated into a CSLO to semi-automate Mueller matrix polarimetry and were used in retinal image acquisition. Success rates of Fourier transform based edge detection filters, designed to improve the registration of retinal images, were compared. The acquired polarimetry images were used to reassess 2 image quality enhancement techniques, Mueller matrix reconstruction (MMR) and Stokes vector reconstruction (SVR), focusing on the role of auto-contrasting or normalization within the techniques and the degree to which auto-contrasting or normalization is responsible for image quality improvement of the resulting images. Mueller matrix polarimetry was also applied to find the retardance image of a malaria infected retinal blood vessel imaged in a confocal scanning laser microscope (CSLM) to visualize hemozoin within the vessel. Image quality enhancement techniques were also applied and image quality improvement was quantified for this blood vessel. The semi-automation of Mueller matrix polarimetry yielded a significant reduction in experimental acquisition time (80%) and a non-significant reduction in registration time (44%). A larger sample size would give higher power and this result might become significant. The reduction in registration time was most likely due to less movement of the eye, particularly in terms of decreased rotation seen between registered images. Fourier transform edge detection methods increased the success rate of registration from 73.9% to 92.3%. Assessment of the 2 MMR images (max entropy and max signal-to-noise ratio (SNR)) showed that comparison to the best CSLO images (not auto-contrasted) yielded significant average image quality improvements of 158% and 4% when quantified with entropy and SNR, respectively. When compared to best auto-contrasted CSLO images, significant image quality improvements were 11% and 5% for entropy and SNR, respectively. Images constructed from auto-contrasted input images were of significantly higher quality than images reconstructed from original images. Of the 2 other images assessed (modified degree of polarization (DOPM) and the first element of the Stokes vector (S0)), DOPM and S0 yielded significant average image quality improvements quantified by entropy except for the DOPM image of the RNFL. SNR was not improved significantly when either SVR image was compared to the best CSLO images. Compared to the best auto-contrasted CSLO images, neither DOPM nor S0 improved average image quality significantly. This result might change with a larger number of participants. When MMR were applied to images of malaria infected retinal slides, image quality was improved by 19.7% and 15.3% in terms of entropy and SNR, respectively, when compared to the best CSLO image. The DOPM image yielded image quality improvements of 8.6% and -24.3% and the S0 image gave improvements of 9.5% and 9.4% in entropy and SNR, respectively. Although percent increase in image quality was reduced when images were compared to initial auto-contrasted CSLO images, the final image quality was improved when auto-contrasting occurred prior to polarimetry calculations for max SNR and max entropy images. Quantitative values of retardance could not be found due to physical constraints in the CSLM that did not allow for characterization of its polarization properties and vibrational noise. Mueller matrix polarimetry used to find the retardance image of a malaria infected retina sample did yield visualization of hemozoin within the vessel but only qualitatively. In conclusion, improvements in the acquisition and registration of CSLO images were successful in leading to considerably shorter experimentation and processing times. In terms of polarimetric image quality improvement techniques, when compared to the best CSLO image. A large proportion of the improvement was in fact due to partially or completely stretching the pixel values across the dynamic range of the images within the algorithm of each technique. However, in general the image quality was still improved by the Mueller matrix reconstruction techniques using both entropy and SNR to generate the CSLO retinal images and the CSLM imaged malaria infected sample. In the malaria sample, retinal blood vessel visualization was also qualitatively improved. The images yielded from Mueller matrix polarimetry applied to a malaria infected retinal sample localized hemozoin within the blood vessel, but a quantitative image of the phase retardance could not be achieved.

The focus of this thesis is the improvement of acquisition and processing of Mueller matrix polarimetry using a confocal scanning laser ophthalmoscope (CSLO) and the application of Mueller matrix polarimetry to image the retina. Stepper motors were incorporated into a CSLO to semi-automate Mueller matrix polarimetry and were used in retinal image acquisition. Success rates of Fourier transform based edge detection filters, designed to improve the registration of retinal images, were compared. The acquired polarimetry images were used to reassess 2 image quality enhancement techniques, Mueller matrix reconstruction (MMR) and Stokes vector reconstruction (SVR), focusing on the role of auto-contrasting or normalization within the techniques and the degree to which auto-contrasting or normalization is responsible for image quality improvement of the resulting images. Mueller matrix polarimetry was also applied to find the retardance image of a malaria infected retinal blood vessel imaged in a confocal scanning laser microscope (CSLM) to visualize hemozoin within the vessel. Image quality enhancement techniques were also applied and image quality improvement was quantified for this blood vessel. The semi-automation of Mueller matrix polarimetry yielded a significant reduction in experimental acquisition time (80%) and a non-significant reduction in registration time (44%). A larger sample size would give higher power and this result might become significant. The reduction in registration time was most likely due to less movement of the eye, particularly in terms of decreased rotation seen between registered images. Fourier transform edge detection methods increased the success rate of registration from 73.9% to 92.3%. Assessment of the 2 MMR images (max entropy and max signal-to-noise ratio (SNR)) showed that comparison to the best CSLO images (not auto-contrasted) yielded significant average image quality improvements of 158% and 4% when quantified with entropy and SNR, respectively. When compared to best auto-contrasted CSLO images, significant image quality improvements were 11% and 5% for entropy and SNR, respectively. Images constructed from auto-contrasted input images were of significantly higher quality than images reconstructed from original images. Of the 2 other images assessed (modified degree of polarization (DOPM) and the first element of the Stokes vector (S0)), DOPM and S0 yielded significant average image quality improvements quantified by entropy except for the DOPM image of the RNFL. SNR was not improved significantly when either SVR image was compared to the best CSLO images. Compared to the best auto-contrasted CSLO images, neither DOPM nor S0 improved average image quality significantly. This result might change with a larger number of participants. When MMR were applied to images of malaria infected retinal slides, image quality was improved by 19.7% and 15.3% in terms of entropy and SNR, respectively, when compared to the best CSLO image. The DOPM image yielded image quality improvements of 8.6% and -24.3% and the S0 image gave improvements of 9.5% and 9.4% in entropy and SNR, respectively. Although percent increase in image quality was reduced when images were compared to initial auto-contrasted CSLO images, the final image quality was improved when auto-contrasting occurred prior to polarimetry calculations for max SNR and max entropy images. Quantitative values of retardance could not be found due to physical constraints in the CSLM that did not allow for characterization of its polarization properties and vibrational noise. Mueller matrix polarimetry used to find the retardance image of a malaria infected retina sample did yield visualization of hemozoin within the vessel but only qualitatively. In conclusion, improvements in the acquisition and registration of CSLO images were successful in leading to considerably shorter experimentation and processing times. In terms of polarimetric image quality improvement techniques, when compared to the best CSLO image. A large proportion of the improvement was in fact due to partially or completely stretching the pixel values across the dynamic range of the images within the algorithm of each technique. However, in general the image quality was still improved by the Mueller matrix reconstruction techniques using both entropy and SNR to generate the CSLO retinal images and the CSLM imaged malaria infected sample. In the malaria sample, retinal blood vessel visualization was also qualitatively improved. The images yielded from Mueller matrix polarimetry applied to a malaria infected retinal sample localized hemozoin within the blood vessel, but a quantitative image of the phase retardance could not be achieved.

This diploma thesis is dealing with the detection of blood vessels pulsation in retinal sequences. The goal is to create an algorithm for objective evaluation of pulsation in retinal video sequences.

Retinal imaging enables the visualization of a portion of the human microvasculature in-vivo and non-invasively. The scanning laser ophthalmoscope (SLO), provides images characterized by an ultra-wide field of view (UWFoV) covering approximately 180-200º in a single scan, minimizing the discomfort for the subject. The microvasculature visible in retinal images and its changes have been vastly investigated as candidate biomarkers for different types of systemic conditions like cardiovascular disease (CVD), which currently remains the main cause of death in Europe. For the CARMEN study, UWFoV SLO images were acquired from more than 1,000 people who were recruited from two studies, TASCFORCE and SCOT-HEART, focused on CVD. This thesis presents an automated system for SLO image processing and computation of candidate biomarkers to be associated with cardiovascular risk and MRI imaging data. A vessel segmentation technique was developed by making use of a bank of multi-scale matched filters and a neural network classifier. The technique was devised to minimize errors in vessel width estimation, in order to ensure the reliability of width measures obtained from the vessel maps. After a step of refinement of the centrelines, a multi-level classification technique was deployed to label all vessel segments as arterioles or venules. The method exploited a set of pixel-level features for local classification and a novel formulation for a graph cut approach to partition consistently the retinal vasculature that was modelled as an undirected graph. Once all the vessels were labelled, a tree representation was adopted for each vessel and its branches to fully automate the process of biomarker extraction. Finally, a set of 75 retinal parameters, including information provided by the periphery of the retina, was created for each image and used for the biomarker investigation.

Abstract Glaucoma is a group of eye diseases characterized by a chronic, progressive optic neuropathy. During the disease process, the axon damage of the retinal ganglion cells leads to changes in the retinal nerve fiber layer, causing optic nerve head, and visual field defects typical of glaucoma. The Heidelberg Retina Tomograph (HRT) is a confocal scanning laser imaging device acquiring and analysing three-dimensional data of the ocular fundus wit good accuracy and reproducibility. Conventional planimetric measurements were compared to those taken with the HRT in a pilot study of 12 eyes with early glaucomatous optic disc, retinal nerve fiber layer and/or visual field abnormalities. The neuroretinal rim area measurements and cup-to-disc area ratio did not differ statistically from each other when using these two different methods. The effect of four different reference levels on the HRT parameter measurement values was tested in two separate studies. In the first study there were 67 eyes, 40 of the eyes were healthy and 27 eyes had glaucoma of different stages. Then, 279 eyes, 180 of which were non-glaucomatous and 99 glaucoma eyes, were included in another study. The flexible reference level gave the most reliable HRT parameter measurement values in both non-glaucomatous and glaucomatous eyes. The ability of the HRT parameters to separate between non-glaucomatous and glaucomatous eyes was tested in 77 eyes, 40 of the eyes were non-glaucomatous, 10 ocular hypertensives and 27 eyes had different stages of glaucoma. The reference level dependent HRT parameters cup-to-disc area ratio, vertical linear cup-to-disc ratio, mean retinal nerve fiber layer thickness (RNFLt) and rim volume as well as the reference level non-dependent HRT parameter, cup shape measure (CSM), separated best between the clinical groups. The best combination of the HRT and other structural and functional parameters in separating between non-glaucomatous and glaucomatous eyes was studied in 55 eyes. There were 32 non-glaucomatous eyes and 23 eyes with ocular hypertension or glaucoma. CSM, RNFLt, together with age- and lens coloration-corrected mean deviation of the B/Y perimetry showed good discrimination (ROC area 0.91) between non-glaucomatous and glaucomatous eyes. / Tiivistelmä Glaukooma koostuu joukosta hitaasti eteneviä näköhermon rappeumasairauksia. Sairausprosessin aikana verkkokalvon gangliosolujen aksonivaurio johtaa muutoksiin verkkokalvon hermosäiekerroksessa ja näköhermon päässä aiheuttaen glaukoomalle tyypillisiä näkökenttämuutoksia. The Heidelberg Retina Tomograph (HRT) on konfokaali laserskanneritekniikkaan perustuva kuvantamislaite, joka tuottaa ja analysoi silmänpohjasta saatua kolmiulotteista mittaustietoa tarkasti ja toistettavasti. Tavanomaisen planimetrian antamia mittaustuloksia verrattiin HRT:n antamiin tuloksiin 12:ssa silmässä, joissa oli todettu varhaisia glaukoomamuutoksia. Näköhermon pään hermoreunan (rim) pinta-ala ja keskuskuopan suhde papillan läpimittaan eivät poikenneet tilastollisesti toisistaan näitä kahta menetelmää käytettäessä. Neljän eri referenssitason vaikutusta HRT-parametrien mittausarvoihin testattiin kahdessa eri tutkimuksessa. Ensimmäisen tutkimusaineisto koostui yhteensä 67:stä silmästä, joista 40 oli terveitä ja 27:ssä eriasteisia glaukoomamuutoksia. Toisessa tutkimuksessa oli yhteensä 279 silmää, joista 180 oli terveitä ja 99:llä oli glaukooma. Papillomakulaarisäikeisiin tukeutuva, fleksiibeli referenssitaso antoi luotettavimmat HRT-parametrien mittaustulokset sekä terveissä että glaukoomasilmissä. HRT-parametrien kykyä erottaa terveet silmät glaukomatoottisista testattiin yhteensä 77:ssä silmässä, joista 40 oli terveitä, 10 oli korkeapaineisia ilman glaukoomamuutoksia, ja 27:ssä oli glaukoomamuutoksia. Referenssitasosta riippuvaiset HRT-parametrit, keskuskuopan suhde papillan läpimittaan, vertikaali-lineaarinen keskuskuopan suhde papillan läpimittaan, keskimääräinen verkkokalvon hermosäiekerroksen paksuus (RNFLt) ja `rim´:in tilavuus samoin kuin referenssitasosta riippumaton keskuskuopan ´vinous´-mitta (CSM) erottelivat parhaiten nämä kliiniset ryhmät toisistaan. Terveitä ja glaukoomasilmiä erottelevaa HRT:n ja muiden rakenteellisten ja toiminnallisten parametrien kombinaatiota etsittiin 55:n silmän aineistosta. Silmistä 32 oli terveitä ja 23 korkeapaineisia ja/tai glaukoomavaurioisia. CSM ja RNFLt, yhdessä iän ja mykiövärjäytymisen suhteen korjatun sinikeltaperimetrian keskipoikkeaman kanssa osoittivat hyvää erottelukykyä (ROC area 0.91) terveiden ja glaukoomasilmien välillä.

Studies demonstrate neuro-retinal damage in patients with diabetes and no clinically visible diabetic retinopathy. It is unknown which retinal regions are most vulnerable to diabetes. We hypothesized that the standard and slow-flash (sf-) multifocal electroretinogram (mfERG) and adaptive optics (AO) imaging will localize retinal regions of vulnerability. Fifty-five adolescents with diabetes and 54 controls underwent mfERG testing to isolate predominately retinal bipolar cell activity and sf-mfERG testing to isolate three oscillatory potentials (OPs) from intraretinal amacrine and interplexiform cells. Greatest mfERG delays were in the superior temporal quadrant and at 5°-10° eccentricity. Greatest sf-mfERG delays were found at different eccentricities for each OP. Twenty adolescents with diabetes and 14 controls underwent AO imaging. No significant differences in cone photoreceptor density were found; however, patients showed a trend towards reduced density in the superior nasal region. Inner retinal structures may be more susceptible to damage by diabetes than outer retinal structures.

Studies demonstrate neuro-retinal damage in patients with diabetes and no clinically visible diabetic retinopathy. It is unknown which retinal regions are most vulnerable to diabetes. We hypothesized that the standard and slow-flash (sf-) multifocal electroretinogram (mfERG) and adaptive optics (AO) imaging will localize retinal regions of vulnerability. Fifty-five adolescents with diabetes and 54 controls underwent mfERG testing to isolate predominately retinal bipolar cell activity and sf-mfERG testing to isolate three oscillatory potentials (OPs) from intraretinal amacrine and interplexiform cells. Greatest mfERG delays were in the superior temporal quadrant and at 5°-10° eccentricity. Greatest sf-mfERG delays were found at different eccentricities for each OP. Twenty adolescents with diabetes and 14 controls underwent AO imaging. No significant differences in cone photoreceptor density were found; however, patients showed a trend towards reduced density in the superior nasal region. Inner retinal structures may be more susceptible to damage by diabetes than outer retinal structures.