Automated

An often used Laplacian filter for enhancing signals at object boundaries and dots. It works with XY, XYZ, XYZ-T, XYZ-T-Ch1, XYZT-C1-C2 images. Distributed as a part of ImageJ plugin FeatureJ, and included in Fiji. The second URL above is the link to its Javadoc. (imagescience.feature.Laplacian). A primer for using this class in Jython script is in CMCI Jython/Fiji cookbook: FeatureJ.

Priism is an image processing, analysis and visualisation application for multidimensional light and electron microscopy data. It uses the Image Visualization Environment (IVE) library, which is a closed-source C library with documented C and FORTRAN APIs. Priism/IVE was developed at UCSF, and can be obtained free of charge by sending an email request to the developers. Many of the same processing and analysis functions available in other image processing applications can be found here (cropping, resizing, spatial filters, Fourier transforms, segmentation etc.), and it includes a comprehensive html-based offline manual. The "SoftWoRx" package of GE Healthcare (formerly Applied Precision) was based on Priism/IVE.

Fluorescence spectroscopy by image correlation is a technique that allows analysing and characterizing the different molecular dynamics from a sequence of fluorescence images. Many image correlation techniques have been developed for different applications but in particular to study the mechanisms of cell adhesion during migration. These techniques can be used with most imaging modalities: e.g. fluorescence widefield, confocal microscopy, TIRFM. They allow to obtain information such as the density in molecules, diffusion coefficients, the presence of several populations, or the direction and speed of a movement corresponding to active transport when spatial and temporal correlations are taken into account (STICS: Spatio-Temporal Image Correlation Spectroscopy).

This plugin is based on ICS_tools plugin by Fitz Elliott, available here.

Some bugs have been removed, ROI does not need to be squared, fitting is weighted in order to give more weight to the smaller lags (temporal or spatial)

Exemple of use on sample data [fluorescent beads](http://biii.info/node/2577 "Beads") - Select an ROI, start by ICS to get the right PSF size - Then run TICS and select diffusion, or diffusion plus flow model. Remove the first line (autocorrelation) which corresponds to the noise autocorrelation before fitting.

The workflow consists of firstly identifying spot (which can be also gravity center of cells identified by another method), and then secondly compute trajectories by linking these spots by global optimisation with a cost function. This method is part of the methods evaluated in Chanouard et al (2014) as "method 9" and is described in detail in its supplementary PDF (page 65).

Dependencies

Following plugins are required.

1. JAR to be placed under IJ plugin directory
2. A pdf file with instructions and output description is also available in the zip .
3. MTrackJ : Used for visualization of tracks. Preinstalled in Fiji.
4. Imagescience.jar: This library is used by MTrackJ. Use update site to install this plugin.
5. jama.jar. Preinstalled in Fiji.

##Advantages:

• support blinking (with a parameters allowing it or not)
• fast,
• can be used in batch, some analysis results provided.
• No dynamic model.
• The tracking part is not dependent of ImageJ.

Pitfalls:

• does not support division
• the optimization algorithm used is a simulated annealing, so results can be slightly different between two runs.
• No Dynamic model (so less good results but can be used for a first study of the kind of movements)

##The sample data

The parameters used for this example data Beads, were

1. detection: 150
2. the max distance in pixels: 20
3. max allowed disappearance in frame: 1

HDF5 is a data format for storing extremely large and complex data collections. This Fiji/ImageJ HDF5 plugin saves and loads 2D - 5D datasets with flexible options.

In Fiji, the plugin is downloadable via update site "HDF5".