ROI

ROIs

PyFRAP allows the user to define arbitrary Regions Of Interest (ROI) to evaluate simulation and analysis results. ROIs in PyFRAP are defined as ROI objects and saved in the embryo objects ROIs list. If you want to learn more about the ROI object and its capabilities, check out the ROI API here .

Different ROIs available

PyFRAP allows the user to define an arbitrary number of ROIs. There are different ROI types available:

• Circular
• Rectangular
• Polygonal

These shapes refer to the plane in which your experiment is conducted (by default the x-y-plane). Each ROI can be either finite or infinte in the remaining direction (by default z).

Moreover, you can combine multiple ROIs be either adding or substracting different ROIs.

The default ROIs

PyFRAP offers the quick creation of a default set of ROIs that were found optimal for FRAP experiments in radial geometries. The default set contains the following ROIs:

• All: ROI covering all pixels and mesh nodes.
• All Square: ROI covering all pixels and mesh nodes inside bleached region.
• All Out: ROI covering all pixels and mesh nodes outside bleached region.
• Slice: ROI covering all pixels and mesh nodes inside recorded field of view. This ROI is the master ROI by default, see also here
• Slice rim: ROI covering all pixels that are not used for rim concentration computation.
• Rim: ROI covering all pixels that are used for rim concentration computation.
• Bleached Square: ROI covering all pixels and mesh nodes inside bleached region and imaging slice.
• Out: ROI covering all pixels and mesh nodes outside bleached region but inside imaging slice.

PyFRAP also offers to create a minimal set of ROIs, that is, only Slice, Bleached Square, Slice rim and Rim. All ROI options including various ROI creation wizards and an ROI manager can be found in the GUI under Embryo -> ROIs.

Creating ROIs

Creating ROIs using the PyFRAP GUI

The PyFRAP GUI offers three different ways to create and edit ROIs:

• Create default ROIs: Simplest way to create a default set of ROIs.
• Use ROI Wizard: Creates the same set of ROIs as default ROIs, but lets you choose their dimensions more exactly.
• Use ROI Manager: Opens the ROI manager. The ROI Manager allows for the creation of arbitrary sets of ROIs.

What is saved in a ROI?

ROIs mainly ontain:

• Image coordinates lying inside the ROI.
• Mesh nodes lying inside the ROI.
• Mean image intensities inside the ROI.
• Mean simulation intensities inside the ROI.
• Parameters describing ROI geometry.

The master ROI

The master ROI is a special ROI in PyFRAP necessary to tell PyFRAP what is actual recorded data and what is "made up" values through rim approximation. Thus generally one selects the ROI describing the actual region of imaging.

Which ROI should I use for fitting?

PyFRAP allows you to use any ROI for fitting simulation results to data analysis results, however not every ROI is a good choice for fitting. A good rule-of-thumb is: The more I know about concentration in a ROI, the more suitable it is for fitting. That is, ROIs having the largest overlapping with the imaging regions in general suit well, while ROIs that have a lot of external pixels or include a lot of mesh nodes that are filled with the rim concentration prove to be bad choices.

In case of of the default ROIs, the ROIs Slice and Bleached Square generally prove to be the best choice, since

• Slice is covering most of the image (if chosen appropiatly), so most of the available data is included into the fitting, and
• Bleached Square generally provides a good recovery curve showing the typical FRAP recovery behaviour.