Hi everyone,
I am a student trying to quantify the number of cells in an image using ImageJ.
Is there a way to bring all my images from a set of experiments to a set intensity?
For example, if I'm doing live cell imaging where the efficacy of my fluorescent dye may decrease by the time I get to my last few plates due to it being in a working solution that is not stable over long periods, or perhaps I want to regulate for the difference in dye activity and intensity per dish.
In my case, I am trying to count the number of cells I see per channel in a given image. Using ImageJ, I have set intensity thresholds, defined size limits, and defined circularity thresholds. However, if one dish has a really weak dye signal, and I can see that there are around 80 to 100 cells in view, but my counter may only pick up 30 to 60, that throws off my count when compared to dishes with more powerful signal intensity.
Is there a way to automatically change the intensity of all images to a standardized value so that they all match the same intensity per channel? I am strictly comparing the number of cells per channel, and not measuring intensity differences at all. I am working with .tiff images, as well as .czi.
Thank you very much for your help and guidance!
You can use FIJI (ImageJ) to batch process all the images to a set intensity value. A simple answer is:
- Open the image in FIJI
- Convert them to a 8 bit image (Image > Type > 8-bit)
- Threshold all the images to a set value (Image > Adjust > Threshold > "select the min and max threshold" > Apply). Set the values until you see the cells of interest in all the images.
- Run your segmentation and parameters to count the cells now on the above images
You can create a macros to make this automated in FIJI. As you have mentioned that you just wanted to count the cells and not the intensity, this should work.
Good luck.
@Sathya
Hi Sathya Srinivasan,
Thank you so much for your help! Will definitely try this out.
Kindest regards,
Fahad
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