Analysis of 2D cell migration in Igor Pro
This is a set of functions to load particle tracks and analyse 2D cell migration in Igor.
- Cells are tracked manually in ImageJ/FIJI using Manual Tracking.
- Save the outputs as csv, or copy-paste them into an Excel workbook*.
- Save a copy of
CellMigration.ipfin Wavemetrics/Igor Pro 7 User Files/User Procedures. Open in Igor and compile.
- Run using Macros>Cell Migration...
The dialog asks the user how many conditions are to be loaded and analysed. At this point, please confirm the time step and pixelsize of the movies.
Next, a panel pops up where the user specifies:
- The name of each condition
- Either the directory containing all the csvs from that condition or the Excel workbook containing the data
- OPTIONALLY, data containing offset information (if stage travel is an issue in the experiments). Again, either as a directory of CSVs or an Excel workbook.
Now click Do it and Igor will do the rest!
It will calculate and display the following:
- Cumulative distance over time
- Instantaneous speed over time, histogram of velocities
- Tracks of all cells for visualisation
- Directionality ratio (d/D) over time
- Mean squared displacement
- Direction autocorrelation
- Cell turning
Reports are made for each condition and also a summary layout comparing all conditions. Select Macros>Save Reports to save all reports as PDF (Mac) or EMF (Windows).
NOTE: no headers in Excel file. Keep data to columns A-H, max of 2000 rows.
- A - 0 - ImageJ row
- B - 1 - Track No
- C - 2 - Slice No
- D - 3 - x (in px)
- E - 4 - y (in px)
- F - 5 - distance
- G - 6 - velocity (speed)
- H - 7 - pixel value
Colours are taken from Paul Tol SRON stylesheet. A maximum of 12 conditions are handled properly, with >12 conditions having non-unique colours. By editing the 3-column colorWave (root:colorWave) it is possible to recolor all the plots by subsequently clicking Macros > Recolor Everything.
Written for IgorPro 7. From v 1.03 there is no back-compatability for IgorPro 6.37.
* CSV output: This is the preferred method. Save the output direct from ImageJ. Use a directory of CSVs percondition. They can be named anything, as long as they have .csv extension. If you need offsetting then the same named files are needed for this (in a different directory).
* Excel: Use 1 workbook per condition. Suggest that each sheet is a field of view, containing all cells in the field. So for two experimental conditions with 10 multipoints each, you will have two workbooks each with 10 worksheets.
* Offsetting: For offsetting data, a directory of CSVs or workbook per condition is needed with corresponding files or sheets to the primary data. It is important that every frame has a tracked point.