Two plus one is almost three: a fast approximation for multi-view deconvolution

Multi-view deconvolution is a powerful image-processing tool for light sheet fluorescence microscopy, providing isotropic resolution and enhancing the image content. However, performing these calculations on large datasets is computationally demanding and time-consuming even on high-end workstations. Especially in long-time measurements on developing animals, huge amounts of image data are acquired. To keep them manageable, redundancies should be removed right after image acquisition. To this end, we report a fast approximation to three-dimensional multi-view deconvolution, denoted 2D+1D multi-view deconvolution, which is able to keep up with the data flow. It first operates on the two dimensions perpendicular and subsequently on the one parallel to the rotation axis, exploiting the rotational symmetry of the point spread function along the rotation axis. We validated our algorithm and evaluated it quantitatively against two-dimensional and three-dimensional multi-view deconvolution using simulated and real image data. 2D+1D multi-view deconvolution takes similar computation time but performs markedly better than the two-dimensional approximation only. Therefore, it will be most useful for image processing in time-critical applications, where the full 3D multi-view deconvolution cannot keep up with the data flow

Mosaicing of Single Plane Illumination Microscopy Images Using Groupwise Registration and Fast Content-Based Image Fusion

Single Plane Illumination Microscopy (SPIM; Huisken et al., Nature 305(5686):1007–1009, 2004) is an emerging microscopic technique that enables live imaging of large biological specimens in their entirety. By imaging the living biological sample from multiple angles SPIM has the potential to achieve isotropic resolution throughout even relatively large biological specimens. For every angle, however, only a relatively shallow section of the specimen is imaged with high resolution, whereas deeper regions appear increasingly blurred. In order to produce a single, uniformly high resolution image, we propose here an image mosaicing algorithm that combines state of the art groupwise image registration for alignment with content-based image fusion to prevent degrading of the fused image due to regional blurring of the input images. For the registration stage, we introduce an application-specific groupwise transformation model that incorporates per-image as well as groupwise transformation parameters. We also propose a new fusion algorithm based on Gaussian filters, which is substantially faster than fusion based on local image entropy. We demonstrate the performance of our mosaicing method on data acquired from living embryos of the fruit fly, Drosophila, using four and eight angle acquisitions

Multiple-view microscopy with light-sheet based fluorescence microscope

The axial resolution of any standard single-lens light microscope is lower than its lateral resolution. The ratio is approximately 3-4 when high numerical aperture objective lenses are used (NA 1.2 -1.4) and more than 10 with low numerical apertures (NA 0.2 and below). In biological imaging, the axial resolution is normally insufficient to resolve subcellular phenomena. Furthermore, parts of the images of opaque specimens are often highly degraded or obscured. Multiple-view fluorescence microscopy overcomes both problems simultaneously by recording multiple images of the same specimen along different directions. The images are digitally fused into a single high-quality image. Multiple-view imaging was developed as an extension to the light-sheet based fluorescence microscope (LSFM), a novel technique that seems to be better suited for multiple-view imaging than any other fluorescence microscopy method to date. In this contribution, the LSFM properties, which are important for multiple-view imaging, are characterized and the implementation of LSFM based multiple-view microscopy is described. The important aspects of multiple-view image alignment and fusion are discussed, the published algorithms are reviewed and original solutions are proposed. The advantages and limitations of multiple-view imaging with LSFM are demonstrated using a number of specimens, which range in size from a single yeast cell to an adult fruit fly and to Medaka fish

The Adaptive Particle Representation (APR) for Simple and Efficient Adaptive Resolution Processing, Storage and Simulations

This thesis presents the Adaptive Particle Representation (APR), a novel adaptive data representation that can be used for general data processing, storage, and simulations. The APR is motivated, and designed, as a replacement representation for pixel images to address computational and memory bottlenecks in processing pipelines for studying spatiotemporal processes in biology using Light-sheet Fluo- rescence Microscopy (LSFM) data. The APR is an adaptive function representation that represents a function in a spatially adaptive way using a set of Particle Cells V and function values stored at particle collocation points P∗. The Particle Cells partition space, and implicitly define a piecewise constant Implied Resolution Function R∗(y) and particle sampling locations. As an adaptive data representation, the APR can be used to provide both computational and memory benefits by aligning the number of Particle Cells and particles with the spatial scales of the function. The APR allows reconstruction of a function value at any location y using any positive weighted combination of particles within a distance of R∗(y). The Particle Cells V are selected such that the error between the reconstruction and the original function, when weighted by a function σ(y), is below a user-set relative error threshold E. We call this the Reconstruction Condition and σ(y) the Local Intensity Scale. σ(y) is motivated by local gain controls in the human visual system, and for LSFM data can be used to account for contrast variations across an image. The APR is formed by satisfying an additional condition on R∗(y); we call the Resolution Bound. The Resolution Bound relates the R∗(y) to a local maximum of the absolute value function derivatives within a distance R∗(y) or y. Given restric- tions on σ(y), satisfaction of the Resolution Bound also guarantees satisfaction of the Reconstruction Condition. In this thesis, we present algorithms and approaches that find the optimal Implied Resolution Function to general problems in the form of the Resolution Bound using Particle Cells using an algorithm we call the Pulling Scheme. Here, optimal means the largest R∗(y) at each location. The Pulling Scheme has worst-case linear complexity in the number of pixels when used to rep- resent images. The approach is general in that the same algorithm can be used for general (α,m)-Reconstruction Conditions, where α denotes the function derivative and m the minimum order of the reconstruction. Further, it can also be combined with anisotropic neighborhoods to provide adaptation in both space and time. The APR can be used with both noise-free and noisy data. For noisy data, the Reconstruction Condition can no longer be guaranteed, but numerical results show an optimal range of relative error E that provides a maximum increase in PSNR over the noisy input data. Further, if it is assumed the Implied Resolution Func- tion satisfies the Resolution Bound, then the APR converges to a biased estimate (constant factor of E), at the optimal statistical rate. The APR continues a long tradition of adaptive data representations and rep- resents a unique trade off between the level of adaptation of the representation and simplicity. Both regarding the APRs structure and its use for processing. Here, we numerically evaluate the adaptation and processing of the APR for use with LSFM data. This is done using both synthetic and LSFM exemplar data. It is concluded from these results that the APR has the correct properties to provide a replacement of pixel images and address bottlenecks in processing for LSFM data. Removal of the bottleneck would be achieved by adapting to spatial, temporal and intensity scale variations in the data. Further, we propose the simple structure of the general APR could provide benefit in areas such as the numerical solution of differential equations, adaptive regression methods, and surface representation for computer graphics