Tracking intracellular vesicles in cells

Hello community,
I need help regarding tracking of intracellular vesicles in cells. I have played around a bit with Particle Tracker 2D/3D included in Mosaic Toolsuite. I think i manage to track particles to some extent. But, i am struggling to get track length and velocity for each particle. I will appreciate any help/advice/suggestion.
I could not upload the multiTIFF image. Here is dropbox link: https://www.dropbox.com/s/s94v6o5i0kxdc3s/cell1.tif?dl=0
I have attached herein the first frame of the file. What I am aiming to do is to track the vesicles (total 60 frames) and obtain track length and velocity of individual particles.
cell1-2.tif (528.1 KB)
Thank you so much for your help.

Hi @sphuyal and welcome to the forum!

I believe you could get good results using the well documented TrackMate in Fiji. I used it on your example cell1 and got some encouraging results concerning spot detection


I think if you follow the documentation you should be able to calculate the features you are looking for

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Thank you so much @CellKai. If it is not too much work, could you please share some of your parameters that you used in TrackMate? It will help me immensely. I am a total novice in tracking and was overwhelmed by the options in Trackmate.

sure, no problem.
The first thing I noticed is that your image scale for “cell1.tiff” seems incorrect and is 0.0000023 micron/px with a frame interval of 0 seconds, so I think you should manually update the spacial calibration of the image. Since I did not know the real values, I set it to 1 micron/px and 1 second/frame by selecting the image and editing its properties in the Image > properties dialog.

Then I started Trackmate and continued next with the below settings:

  • select a detector: Log detector

  • select a view: Hyperstack Displayer
  • Set filtes on spots: quality above 9.38 AND mean intensity below 420.34
  • Select a tracker: simple LAP tracker
    • here it would be important for you to consult the documentation on what “linking max distance”, “gap-closing max distance” and “Gap-closing max frame gap” refer to so you can choose meaningful values for your experiment. I just used the default values.

Now the tracks are created and you can set for example a couple of filters concerning their velocity or duration.
in the next menu you have the option to view all track statistics by pressing “Analysis”

.

the final menu lets you export the spot statistics or all tracks and also lets you capture an overlay, i.e. a movie of your spots following the traces.

Hope this helps!

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Thank you very much @CellKai! I am truly grateful to you for this support!