A Patch-Based Method for Repetitive and Transient Event Detection in Fluorescence Imaging

Automatic detection and characterization of molecular behavior in large data sets obtained by fast imaging in advanced light microscopy become key issues to decipher the dynamic architectures and their coordination in the living cell. Automatic quantification of the number of sudden and transient events observed in fluorescence microscopy is discussed in this paper. We propose a calibrated method based on the comparison of image patches expected to distinguish sudden appearing/vanishing fluorescent spots from other motion behaviors such as lateral movements. We analyze the performances of two statistical control procedures and compare the proposed approach to a frame difference approach using the same controls on a benchmark of synthetic image sequences. We have then selected a molecular model related to membrane trafficking and considered real image sequences obtained in cells stably expressing an endocytic-recycling trans-membrane protein, the Langerin-YFP, for validation. With this model, we targeted the efficient detection of fast and transient local fluorescence concentration arising in image sequences from a data base provided by two different microscopy modalities, wide field (WF) video microscopy using maximum intensity projection along the axial direction and total internal reflection fluorescence microscopy. Finally, the proposed detection method is briefly used to statistically explore the effect of several perturbations on the rate of transient events detected on the pilot biological model

An extended model of vesicle fusion at the plasma membrane to estimate protein lateral diffusion from TIRF microscopy images.

BACKGROUND: Characterizing membrane dynamics is a key issue to understand cell exchanges with the extra-cellular medium. Total internal reflection fluorescence microscopy (TIRFM) is well suited to focus on the late steps of exocytosis at the plasma membrane. However, it is still a challenging task to quantify (lateral) diffusion and estimate local dynamics of proteins. RESULTS: A new model was introduced to represent the behavior of cargo transmembrane proteins during the vesicle fusion to the plasma membrane at the end of the exocytosis process. Two biophysical parameters, the diffusion coefficient and the release rate parameter, are automatically estimated from TIRFM image sequences, to account for both the lateral diffusion of molecules at the membrane and the continuous release of the proteins from the vesicle to the plasma membrane. Quantitative evaluation on 300 realistic computer-generated image sequences demonstrated the efficiency and accuracy of the method. The application of our method on 16 real TIRFM image sequences additionally revealed differences in the dynamic behavior of Transferrin Receptor (TfR) and Langerin proteins. CONCLUSION: An automated method has been designed to simultaneously estimate the diffusion coefficient and the release rate for each individual vesicle fusion event at the plasma membrane in TIRFM image sequences. It can be exploited for further deciphering cell membrane dynamics

BioImage-IT: Design data analysis workflows using tools from different languages and keep track of the metadata

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Non-parametric regression for patch-based fluorescence microscopy image sequence denoising

We present a non-parametric regression method for denoising 3D image sequences acquired in fluorescence microscopy. The proposed method exploits 3D+time information to improve the signal-to-noise ratio of images corrupted by mixed Poisson-Gaussian noise. A variance stabilization transform is first applied to the image-data to introduce independence between the mean and variance. This pre-processing requires the knowledge of parameters related to the acquisition system, also estimated in our approach. In a second step, we propose an original statistical patch-based framework for noise reduction and preservation of space-time discontinuities. In our study, discontinuities are related to small moving spots with high velocity observed in fluorescence video-microscopy. The idea is to minimize an objective nonlocal energy functional involving spatio-temporal image patches. The minimizer has a simple form and is defined as the weighted average of input data taken in spatially-varying neighborhoods. The size of each neighborhood is optimized to improve the performance of the pointwise estimator. The performance of the algorithm which requires no motion estimation, is then demonstrated on both synthetic and real image sequences using qualitative and quantitative criteria

Patch-Based Markov Models for Event Detection in Fluorescence Bioimaging

International audienceThe study of protein dynamics is essential for understanding the multi-molecular complexes at subcellular levels. Fluorescent Protein (XFP)-tagging and time-lapse fluorescence microscopy enable to observe molecular dynamics and interactions in live cells, unraveling the live states of the matter. Original image analysis methods are then required to process challenging 2D or 3D image sequences. Recently, tracking methods that estimate the whole trajectories of moving objects have been successfully developed. In this paper, we address rather the detection of meaningful events in spatio-temporal fluorescence image sequences, such as apparent stable "stocking areas" involved in membrane transport. We propose an original patch-based Markov modeling to detect spatial irregularities in fluorescence images with low false alarm rates. This approach has been developed for real image sequences of cells expressing XFP-tagged Rab proteins, known to regulate membrane trafficking

Multiscale neighborhood-wise decision fusion for redundancy detection in image pairs

SIAM Journal Multiscale Modeling & SimulationTo develop better image change detection algorithms, new models able to capture spatio-temporal regularities and geometries present in an image pair are needed. In this paper, we propose a multiscale formulation for modeling semi-local inter-image interactions and detecting local or regional changes in an image pair. By introducing dissimilarity measures to compare patches and binary local decisions, we design collaborative decision rules that use the total number of detections obtained from the neighboring pixels, for different patch sizes. We study the statistical properties of the non-parametric detection approach that guarantees small probabilities of false alarms. Experimental results on several applications demonstrate that the detection algorithm (with no optical flow computation) performs well at detecting occlusions and meaningful changes for a variety of illumination conditions and signal-to-noise ratios. The number of control parameters of the algorithm is small and the adjustment is intuitive in most cases

Sélection d'échelle automatique précise et seuillage localement adapté pour la segmentation de vésicules en microscopie TIRF

National audienceEn imagerie cellulaire et microscopie de fluorescence, la première phase de l'analyse consiste généralement à détecter une grande quantité d'éléments (molécules, vésicules). De nombreuses méthodes de détection ont été développées dans ce contexte, mais elles réclament souvent un paramétrage assez fin et ne peuvent pas toujours s'adapter à l'échelle des éléments à détecter. Nous proposons une méthode incluant une sélection d'échelle automatique précise et un seuillage localement adapté. Des expérimentations quantitatives sur images simulées montrent les avantages et les meilleures performances de notre méthode. Des résultats sur séquences réelles confirment son intérêt

Estimation of the flow of particles within a partition of the image domain in fluorescence video-microscopy

International audienceAutomatic analysis of the dynamic content in fluorescence video-microscopy is crucial for understanding molecular mechanisms involved in cell functions. In this paper, we propose an original approach for analyzing particle trafficking in these sequences. Instead of individually tracking every particle, we estimate the particle flows between predefined regions. This approach allows us to process image sequences with a high number of particles and a low frame rate. We investigate several ways to estimate the particle flow at the cellular level and evaluate their performance in synthetic and real image sequences