Suggestions for removing brightfield illumination artifacts

Hello all,

I have been trying to use the correct illumination calculate/apply modules but have not had any luck. I have tried a number of things and looked at the tutorial but have had little success. I was hoping I could get some input on setting the parameters. I have different time series with 6 time points, and each series will need a different correction. I’ve attached an example time lapse below. You will see the circular artifacts scattered throughout the images that persist throughout the time lapse. They have been causing some issues during segmentation so I’d like to remove them if possible!

link to time series (only 24MB)

I look forward to your feedback!

David

It maybe won’t get rid of every single one, but I’d think inverting your image and then using “apply threshold” to create a mask that keeps only your cells would be a good place to start; the mask could then be applied to your downstream images .

Thanks for the suggestion!

When applying the threshold I was running into the same problems, picking up those illumination artifacts and some background, so that mask wasn’t too helpful. But it gave me a good jumping off point.

Instead of applying a threshold, I used the identify primary objects module with a pretty stringent threshold to avoid any unwanted signal, then I used the expandorshrinkobjects module to expand the objects by 30 pixels. This recovered any actual signal I lost from the stringent threshold, without restoring any of the background. Using the expanded objects as a mask for my downstream identify primary objects was really successful, and allowed me to use a really lenient threshold to make sure that I segmented all of my cells.

Again, thanks for the suggestion!

David

Glad you got something figured out! :+1: