Quantitative measurement of blood velocity in zebrafish with optical vector field tomography.

Microscopy techniques can readily visualize the finest details of embryo vasculature, but still lack to provide a complete three-dimensional representation of blood flow parameters. We present an in-vivo 3D imaging technique, able to reconstruct the blood cell velocity vector over a large volume of zebrafish embryos. This low cost and relatively simple technique is exploited to quantitatively assess blood velocity in the zebrafish tail at different stages of development.

Calcium dynamics in root cells of Arabidopsis thaliana visualized with selective plane illumination microscopy.

Selective Plane Illumination Microscopy (SPIM) is an imaging technique particularly suited for long term in-vivo analysis of transparent specimens, able to visualize small organs or entire organisms, at cellular and eventually even subcellular resolution. Here we report the application of SPIM in Calcium imaging based on Förster Resonance Energy Transfer (FRET). Transgenic Arabidopsis plants expressing the genetically encoded-FRET-based Ca(2+) probe Cameleon, in the cytosol or nucleus, were used to demonstrate that SPIM enables ratiometric fluorescence imaging at high spatial and temporal resolution, both at tissue and single cell level. The SPIM-FRET technique enabled us to follow nuclear and cytosolic Ca(2+) dynamics in Arabidopsis root tip cells, deep inside the organ, in response to different stimuli. A relevant physiological phenomenon, namely Ca(2+) signal percolation, predicted in previous studies, has been directly visualized.

Time-gated optical projection tomography allows visualization of adult zebrafish internal structures.

Optical imaging through biological samples is compromised by tissue scattering and currently various approaches aim to overcome this limitation. In this paper we demonstrate that an all optical technique, based on non-linear upconversion of infrared ultrashort laser pulses and on multiple view acquisition, allows the reduction of scattering effects in tomographic imaging. This technique, namely Time-Gated Optical Projection Tomography (TGOPT), is used to reconstruct three dimensionally the internal structure of adult zebrafish without staining or clearing agents. This method extends the use of Optical Projection Tomography to optically diffusive samples yielding reconstructions with reduced artifacts, increased contrast and improved resolution with respect to those obtained with non-gated techniques. The paper shows that TGOPT is particularly suited for imaging the skeletal system and nervous structures of adult zebrafish.

In vivo label-free three-dimensional imaging of zebrafish vasculature with optical projection tomography.

We introduce flow optical projection tomography, an imaging technique capable of visualizing the vasculature of living specimens in 3-D. The method detects the movement of cells in the bloodstream and creates flow maps using a motion-analysis procedure. Then, flow maps obtained from projection taken at several angles are used to reconstruct sections of the circulatory system of the specimen. We therefore demonstrate an in vivo, 3-D optical imaging technique that, without the use of any labeling, is able to reconstruct and visualize the vascular network of transparent and weakly scattering living specimens.

Angularly selective mesoscopic tomography.

We report three-dimensional tomographic reconstruction of optical parameters for the mesoscopic light-scattering regime from experimentally obtained datasets by employing angularly selective data acquisition. The approach is based on the assumption that the transport coefficient of a scattering medium differs by an order of magnitude for weakly and highly scattering regions. Datasets were obtained by imaging a weakly scattering phantom, which embeds a highly scattering cylinder of two to three photons' mean path length in diameter containing light-absorbing inclusions. Reconstruction results are presented and discussed.