The ROI module is designed to show a trace of the tracked ends and allows the user to select regions of interest (ROIs) from the traces i.e. growth or shrinkage events. It also has two built-in filter functions, which allow the user to discard frames in which the tracker performed poorly, either by using the maximum velocity filter and/or a tracking precision cut-off. For quality monitoring it also creates the kymograph of the microtubule from the movie and shows an overlay of the trace and the kymograph. The selected regions of interest are then written to a file and can further be analysed using the analysis module. The local velocity of the selected trace is calculated using a linear least square fit and the selction is shown in a window. If the user wants to select a specific object he can enter the object ID and the chosen trace will be displayed. Note that each of the modules produces an output file that can either be used within the later module or with any other analysis software, such as Matlab. This allows a dynamic use of the program, as one can separate tracking and analysis.
| Parameter | Description |
|---|---|
| Frame rate | time interval between frames of a stack in ms |
| Pixel size | in nm |
| Specific object | id of tracking object, as determined by tracker |
| Data file | tracker output file |
| image file | original image/ stack of images |
| speed filter | Boolean, cut out points that are too distant from previous |
| maximal speed | cut-off value for speed filter. |
| precision filter | Boolean, filter points that have a large tracking error |
| precision cut off | input for precision filter, filter points with larger error |
| Show kymo | shows a kymograph of the object. Needs image file. |
| Dual color | images containing two color layers |
If you want to solely use the ROI or analysis module on data generated with any other program, read: ref:reader-label
All parameters can be specified in the terminal, by substituting a space ” ” with an underscore “_”.
python image_analyser.py STOP=1 GUI=1 ../Test/movie.nd2
.TIF and .nd2 (version 4.0) are supported at this time. The .TIF support is thanks to tifffile.py, provided by Christoph Gohlke. http://www.lfd.uci.edu/~gohlke/code/tifffile.py.html