FIJI (ImageJ): Tracking Cells, Single Particles or Spot-like Objects with TrackMate and MTrackJ
ฝัง
- เผยแพร่เมื่อ 30 พ.ค. 2024
- Learn how to use FIJI (ImageJ) to track and measure track statistics of moving objects (cells, single particles, spot-like objects) in image sequences. Two (2) methods are demonstrated: automatic tracking with TrackMate and manual tracking (when automatic methods fail) with MTrackJ. See timeline below:
00:00 Introduction
00:42 Auto Tracking with TrackMate
05:03 Manual Tracking with MTrackJ
Acknowledgments:
TrackMate: Jean-Yves Tinevez, Nick Perry, Johannes Schindelin, Genevieve M. Hoopes, Gregory D. Reynolds, Emmanuel Laplantine, Sebastian Y. Bednarek, Spencer L. Shorte, Kevin W. Eliceiri, TrackMate: An open and extensible platform for single-particle tracking, Methods, Available online 3 October 2016, ISSN 1046-2023, dx.doi.org/10.1016/j.ymeth.201.... (www.sciencedirect.com/science/...)
MTrackJ: E. Meijering, O. Dzyubachyk, I. Smal
Methods for Cell and Particle Tracking
Methods in Enzymology, vol. 504, February 2012, pp. 183-200
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![FIJI (ImageJ): Kymographs [Tracking Moving Objects]](http://i.ytimg.com/vi/n9iKjD6HZuw/mqdefault.jpg)
![FIJI (ImageJ): Kymographs [Tracking Moving Objects]](/img/tr.png)
![ทททททททท #27 [หลุยส์ เค โตมายังไง ณ สระบุรี]](http://i.ytimg.com/vi/CaKoYw_Xmtg/mqdefault.jpg)
Thank you so much for the tutorial, it is really helpful.🥳
I’m glad to help. Thanks for watching.
Thank you for your effort in making this video and sharing the analysis methods. I Liked and subscribed your youtube channel.
Thanks for your support, @oldyoungboy0227.
Thank you very much for your great efforts 🌹🌹🌹🌹
Hi Hamza. Thanks for dropping by.
Thank you - really useful!!!
Thanks for watching and following my tutorial series, ELM.
Thank you!!
Hello. Thank you for this video is very helpful. I was wondering what format do you use to analyse using TrackMate? Thanks
Hi @Marisol. Thanks for your kind feedback. I don't quite understand what you mean by format...is it the detector (e.g. DoG, LoG, etc.)?
@@johanna.m.dela-cruz sorry, I should have phrased my question better. When you import the file to analyse to Fiji, do you import it as an .avi file or as a stack of .tiff files? Thank you!
@Marisol Pérez Toledo You should load a stack of images. Tiffs are fine.
hello, thanks for the video. my DoG calculator is asking me for the estimated size in pixels, while yours says microns. is there way to change this?
also can we measure the velocity of the particles through the Manual tracking method using MTrack J plugin?
Hi @Kevin TRQ. If your image is spatially calibrated (for example, in microns, nm or mm), the DoG detector will know what units your image has. If it is not calibrated, then you will have to set the scaling of your images.
For manual tracking with MTrackJ, after you have created the tracks, click on Measure on the GUI. One of the measurements in the Results table is velocity.
@@johanna.m.dela-cruz thanks a lot!🙏💯
Hi Thank you for this awesome video. I have a question how can I calculate my x/y calibration.? and z calibartion
Hi @pac0 mp. I appreciate your nice feedback.
If your image is not calibrated (in pixels), you would need to get information about pixel size from whatever microscope you used. If your files are saved in the microscope manufacturer's format, it is more likely that this information is saved in the metadata of your image. Have you checked out this video on calibrating your images? th-cam.com/video/MCasWMg8z5E/w-d-xo.html
Hi I do not understand where you read the object's size as 5 microns. I am looking at the imageJ bar and I see X, Y, and Z coordinates but not in microns. I see where is says length=4.8 but no unit. Is that the object size microns?
Hi @sarahbeauvais8573. 5 um is an estimated object size. The length of 4.8 is in microns, since the image is calibrated. I just rounded up 4.8.
You are a life saver
Happy to help. Thanks for watching.
thank you for your wonderful work, can you make a video to show how to track the rod-shaped cells? I tried trackmate but failed to track the rod-shaped cells. I am very appreciate that if you could help me.
Hi @joy ye. I’ll see what I can do. I would love to be able to help you.
@@johanna.m.dela-cruz if you need, I could offer the videos for you to try. In my videos, there are two species , one is rod-shaped cells with bigger size and moving with high speed, another is spot-like immobile cells like clustering.
@@joyye2316 that would be helpful.
@@johanna.m.dela-cruz Cool, then can i have your email please? I will send the videos to your email.
@joy ye, you can find it in my channel (“About” section).
hello, when i go in the plugin like you did at 0:45 it doesn't show me a tracking list
do i have to download something else to be able to do that
please
Hi @mrp3248. You will need Fiji, rather than the original version of ImageJ. You can download Fiji at fiji.sc/.
@@johanna.m.dela-cruz thanks, it worked, thank you very much , i'm saved lol
thanks for responding so fast
Hello Johanna,
Thank you very much for your video. I have a photo lapse as samples but I don't know how to import it. I read you said to import it as a stack of images in format .tiffs but I don't know how to create a stack of images. Could you help me please?
Thank youu very much!
Best,
Lorenzo
Hi @Lorenzo Cozzolino. You can try opening your file as a virtual stack. There are several built-in commands in Fiji/ImageJ that you could use: File>Import>Image Sequence;
File>Import>Raw; File>Import>Stack From List;
File>Import>AVI.
Hope this helps.
@@johanna.m.dela-cruz Thank you so much for the quick reply. Gonna try this way :)
@@lorenzocozzolino3020 I got a notification of another question from you, but I can't seem to see it anymore. I'm wondering what kind of file you have that you can't open it as a stack.
can we use this for cell migrantion timelapse image stacks?
Hi @Rubal Singh. Thanks for your interest. Yes, my image sample is actually a time-lapse stack. You will probably have to track your cells (maybe the nuclei) using a different tracking detector (perhaps LoG). Let me know how it goes with your images.
@@johanna.m.dela-cruz OK THANKS
Can we calculate the velocity of these moving objects from the data which we obtained from Tackmate?
Hi. Among the track analyzers built into Trackmate are velocities generated from link speeds between spots (distance between 2 spots divided by the time difference). Mean velocity is the mean of the link velocity over all the links of the track. Maximal, mimimal and median velocity, and Velocity standard deviation are feature values that take the link velocities of a track as a distribution they summarize.
As for velocity of particles, TrackMate does not directly give this. You might have to import the tracking results and compute this yourself.
@@johanna.m.dela-cruz Thank you for your suggestion. I'm not getting things clearly. More clearly, I would like to tell you my problem is that I want to calculate the speed/velocity of lysosomes inside the cell. If you have any suggestions please do let me know to solve this issue.
@@AbdulSalam-ds3qh On the TrackMate window, under Display Options, click the Tracks button (at the bottom part). On the left side, you will find Spots, Edges and Tracks. Click on Tracks. One of the columns will give you the mean speed of each of the tracks found.
How to batch analyze files in a folder using Trackmate
Hi Jamila. I think that you will have to rely on scripting to do batch analysis with TrackMate.
You might have to adapt the script according to what your first image needs. More details can be found here: imagej.net/plugins/trackmate/scripting/scripting
Can we calculate the number/rate of mitochondrial fission？ THANK YOU!
Hello. TrackMate can be set to detect and deal with splitting events like fission (or merging events like fusion). These events are mapped in TrackScheme. I believe the Track Analysis does include speed and the number of split events.
Thank you very much for your reply, I have difficulty with the operation, I use TrackMate's Mask detector, but I can't get the results I want, can you share how to set it up.
@@user-lt6tx9yo9d Perhaps you can send me an email?
@@johanna.m.dela-cruz Thank you very much！I'll email you.
@Matias Gonzalez. The pleasure is all mine. Thanks for watching.