Tracking ROI/spots over time with TrackMate and generate mean intensity

I have B/W videos in which I need to track the brightness of a certain spot over time. That spot might move.
The movements are small, so I think TrackMate with manual tracking would be ideal.
Ideally I would define a few key spots in TrackMate, and all intermediate frames would be calculated by TrackMate (assuming within two key spots at different times, the spot would move there in a linear fashion). Defining 3 spots in a 100 frame video, I would get 100 data points of brightness.

But that’s not possible. If you go manual with TrackMate, you need to define all 100 spots yourself. You’ll never get more data than for the exact spots/frames you defined. How can I circumvent that? “TrackMate tools”, which automatically tries to look for any given spot in the subsequent frames, is useless, as it’s movements do not make any sense.

Also, TrackMate does not allow for a default radius size to be defined. I can’t spend hours just resizing radius sizes for each spot individually just for a few hundred frames in a single video.

Any ideas?

Hi @onetimepost,

in order to use Trackmate for that, you need to set the gap closing distance to a high value so that it connects the spare spots to tracks:

Afterwards, in the very final step of Trackmate, you can generate spots in the gaps to read out the intensity during intermediate time points:

We introduced that at some point to do FRAP analysis using Trackmate, where the read-out spot also disappears for some frames.

I hope that helps! Let us know if it works :wink: