Multiple Object Tracking in Light Microscopy Images Using Graph-based and Deep Learning Methods

Multi-Objekt-Tracking (MOT) ist ein Problem der Bildanalyse, welches die Lokalisierung und Verknüpfung von Objekten in einer Bildsequenz über die Zeit umfasst, mit zahlreichen Anwendungen in Bereichen wie autonomes Fahren, Robotik oder Überwachung. Neben technischen Anwendungsgebieten besteht auch ein großer Bedarf an MOT in biomedizinischen Anwendungen. So können beispielsweise Experimente, die mittels Lichtmikroskopie über mehrere Stunden oder Tage hinweg erfasst wurden, Hunderte oder sogar Tausende von ähnlich aussehenden Objekten enthalten, was eine manuelle Analyse unmöglich macht. Um jedoch zuverlässige Schlussfolgerungen aus den verfolgten Objekten abzuleiten, ist eine hohe Qualität der prädizierten Trajektorien erforderlich. Daher werden domänenspezifische MOT-Ansätze benötigt, die in der Lage sind, die Besonderheiten von lichtmikroskopischen Daten zu berücksichtigen. In dieser Arbeit werden daher zwei neuartige Methoden für das MOT-Problem in Lichtmikroskopie-Bildern erarbeitet sowie Ansätze zum Vergleich der Tracking-Methoden vorgestellt. Um die Performanz der Tracking-Methode von der Qualität der Segmentierung zu unterscheiden, wird ein Ansatz vorgeschlagen, der es ermöglicht die Tracking-Methode getrennt von der Segmentierung zu analysieren, was auch eine Untersuchung der Robustheit von Tracking-Methoden gegeben verschlechterter Segmentierungsdaten erlaubt. Des Weiteren wird eine graphbasierte Tracking-Methode vorgeschlagen, welche eine Brücke zwischen einfach anzuwendenden, aber weniger performanten Tracking-Methoden und performanten Tracking-Methoden mit vielen schwer einstellbaren Parametern schlägt. Die vorgeschlagene Tracking-Methode hat nur wenige manuell einstellbare Parameter und ist einfach auf 2D- und 3D-Datensätze anwendbar. Durch die Modellierung von Vorwissen über die Form des Tracking-Graphen ist die vorgeschlagene Tracking-Methode außerdem in der Lage, bestimmte Arten von Segmentierungsfehlern automatisch zu korrigieren. Darüber hinaus wird ein auf Deep Learning basierender Ansatz vorgeschlagen, der die Aufgabe der Instanzsegmentierung und Objektverfolgung gleichzeitig in einem einzigen neuronalen Netzwerk erlernt. Außerdem lernt der vorgeschlagene Ansatz Repräsentationen zu prädizieren, die für den Menschen verständlich sind. Um die Performanz der beiden vorgeschlagenen Tracking-Methoden im Vergleich zu anderen aktuellen, domänenspezifischen Tracking-Ansätzen zu zeigen, werden sie auf einen domänenspezifischen Benchmark angewendet. Darüber hinaus werden weitere Bewertungskriterien für Tracking-Methoden eingeführt, welche zum Vergleich der beiden vorgeschlagenen Tracking-Methoden herangezogen werden

Deep Learning for Detection and Segmentation in High-Content Microscopy Images

High-content microscopy led to many advances in biology and medicine. This fast emerging technology is transforming cell biology into a big data driven science. Computer vision methods are used to automate the analysis of microscopy image data. In recent years, deep learning became popular and had major success in computer vision. Most of the available methods are developed to process natural images. Compared to natural images, microscopy images pose domain specific challenges such as small training datasets, clustered objects, and class imbalance. In this thesis, new deep learning methods for object detection and cell segmentation in microscopy images are introduced. For particle detection in fluorescence microscopy images, a deep learning method based on a domain-adapted Deconvolution Network is presented. In addition, a method for mitotic cell detection in heterogeneous histopathology images is proposed, which combines a deep residual network with Hough voting. The method is used for grading of whole-slide histology images of breast carcinoma. Moreover, a method for both particle detection and cell detection based on object centroids is introduced, which is trainable end-to-end. It comprises a novel Centroid Proposal Network, a layer for ensembling detection hypotheses over image scales and anchors, an anchor regularization scheme which favours prior anchors over regressed locations, and an improved algorithm for Non-Maximum Suppression. Furthermore, a novel loss function based on Normalized Mutual Information is proposed which can cope with strong class imbalance and is derived within a Bayesian framework. For cell segmentation, a deep neural network with increased receptive field to capture rich semantic information is introduced. Moreover, a deep neural network which combines both paradigms of multi-scale feature aggregation of Convolutional Neural Networks and iterative refinement of Recurrent Neural Networks is proposed. To increase the robustness of the training and improve segmentation, a novel focal loss function is presented. In addition, a framework for black-box hyperparameter optimization for biomedical image analysis pipelines is proposed. The framework has a modular architecture that separates hyperparameter sampling and hyperparameter optimization. A visualization of the loss function based on infimum projections is suggested to obtain further insights into the optimization problem. Also, a transfer learning approach is presented, which uses only one color channel for pre-training and performs fine-tuning on more color channels. Furthermore, an approach for unsupervised domain adaptation for histopathological slides is presented. Finally, Galaxy Image Analysis is presented, a platform for web-based microscopy image analysis. Galaxy Image Analysis workflows for cell segmentation in cell cultures, particle detection in mice brain tissue, and MALDI/H&E image registration have been developed. The proposed methods were applied to challenging synthetic as well as real microscopy image data from various microscopy modalities. It turned out that the proposed methods yield state-of-the-art or improved results. The methods were benchmarked in international image analysis challenges and used in various cooperation projects with biomedical researchers

Psr1p interacts with SUN/sad1p and EB1/mal3p to establish the bipolar spindle

Regular Abstracts - Sunday Poster Presentations: no. 382During mitosis, interpolar microtubules from two spindle pole bodies (SPBs) interdigitate to create an antiparallel microtubule array for accommodating numerous regulatory proteins. Among these proteins, the kinesin-5 cut7p/Eg5 is the key player responsible for sliding apart antiparallel microtubules and thus helps in establishing the bipolar spindle. At the onset of mitosis, two SPBs are adjacent to one another with most microtubules running nearly parallel toward the nuclear envelope, creating an unfavorable microtubule configuration for the kinesin-5 kinesins. Therefore, how the cell organizes the antiparallel microtubule array in the first place at mitotic onset remains enigmatic. Here, we show that a novel protein psrp1p localizes to the SPB and plays a key role in organizing the antiparallel microtubule array. The absence of psr1+ leads to a transient monopolar spindle and massive chromosome loss. Further functional characterization demonstrates that psr1p is recruited to the SPB through interaction with the conserved SUN protein sad1p and that psr1p physically interacts with the conserved microtubule plus tip protein mal3p/EB1. These results suggest a model that psr1p serves as a linking protein between sad1p/SUN and mal3p/EB1 to allow microtubule plus ends to be coupled to the SPBs for organization of an antiparallel microtubule array. Thus, we conclude that psr1p is involved in organizing the antiparallel microtubule array in the first place at mitosis onset by interaction with SUN/sad1p and EB1/mal3p, thereby establishing the bipolar spindle.postprin