Marking and categorizing oocytes

08-may-2025 

This project allows automatic and manual measurements of oocyte cells.

Resulting parameters are cell size and elliptical aspect ratio. Size

is visualized by a red line indicating the diameter of equivalent circle with same area.

A category (0..9) can be added manually, and is visualized by the "hand of a clock".

You can get statistics of any subset of objects that meets a combination of properties

A - Install ObjectJ plugin

-      If not done yet, install objectj_.jar from https://sils.fnwi.uva.nl/bcb/objectj

-      Move objectj_.jar into the ImageJ plugins folder by

       dragging it into the ImageJ main window- so it will be saved in the correct place

-      optionally, choose menu Plugins>ObjectJ

-      Now the menu "ObjectJ" should appear between "Analyze" and "Plugins"

-      If already installed, ObjectJ can be upgraded via: menu ObjectJ>Project>Update ObjectJ

B - Download Sample Images and .ojj project file

-      Download  "Oocyte-Samples" containing two sample images

-      Download project file (currently 'Oocytes-35d.ojj') and move it into the project folder " Oocyte-Samples "

-      Drag the .ojj file into the ImageJ main window to

open this project, wait until the project window

appears and shows the panel for "Linked Images"

   (Alternatively, choose menu ObjectJ>Open Project..).

-      Drag&drop the image files to be measured from the project folder

       into the panel 'Linked Images', or click button 'Link All'

C - Usage

1. Automatic marking of cells

     -  choose menu ObjectJ>Mark Oocytes...

     - In the dialog, if ‘Threshold’ is set to zero, you are asked to

        threshold each image interactively, 1= automatic and >1 uses that value.

2.  Categorize any marked cell

-   Locate the cursor above a marked cell and press

         key 0..9 for category. Cell will obtain that category as hand of a clock,

     e.g. 4 o'clock means category 4.

3.  Kill a marked cell

     Position cursor above a cell an press key ‘K’.

     Alternatively, activate in ObjectJ>Tools the ‘pistol tool’.

4. Mark an elliptical cell manually:

         ObjectJ tool "V" nust be activated ( press key ‘V’ once or twice)-So that

     Click and drag along a line A-B to follow the cell’s major axis.

     After releasing the mouse button, a circular roi is shown.

     Now move the cursor back towards the cell’s center to decrease the ellipse’s minor axis

     Press key 0..9 to assign a category to the elliptical roi

5.  Mark a cell with fixed diameter

     Make the floating circle visible above a small cell (if necessary press key V once or twice),

     Press key 0..9 to define thecategory:

     A line of fixed length (here 185 um at 45 deg) is used to mark the existence of such a cell.

     Cell will be categorized like described above

Floating circle before (left) and after categorizing with key "5" (right)

6. Show/Hide Ellipses [E]

     Shows or hides the elliptical contours of marked cells.

7.  Categorize Uncategorized cells...

     You can set a default category to all objects that have no category yet.

     This is done in all linked images.

     Choose ObjectJ>Categorize all uncategorized cells...

         Note: you cannot automatically undo this action!

8.  Results

     Choose ObjectJ>Show Project Results

     to display numeric results. The following columns are implemented:

     Dia          equivalent diameter in um

     Cat          category 0..9, or -1 if uncategorized, or empty for fixed diameter

     Roundn.  same as Minor/Major (range 0..1)

     Ellipt.      ellipticity= ellipse perimeter/actual perimeter

                   (range 0..1, or empty for manual marking)

     Minor      minor axis (um)

     Major      major axis (um)

     Area3      area (unit is 10^3 um^2 for convenience)

9.     Saving

Periodically save the colored markers via menu ObjectJ>Save project.

(Note that menu "File>Save" only would save an image, not the markers)

10.   Mark your own images

-  Once the described sample project works satisfactory, you can save an empty copy

    of the project file via ObjectJ>Project File> Save an Empty Copy.

- The empty copy contains all internal structure and macros, but no markers or linked images.

-  Link your images via ObjectJ>Linked Images> Link Image from project Folder

- Make sure your linked images are scaled correctly (eg pixels per um)

11.   Notes

- cells with fixed diameters don't have any quantitative result entries.

- marking a cell with "fixed diameter" does not invite to add a category

- Categorize all unmarked cells does not  categorize those with fixed diameter