Note: I'm using the Carl Zeiss Zen 2 software, and we're not licensed for the image analysis module.
I have double-labeled cells (EdU and DAPI) and in addition to counting nuclei, I was hoping to extract the intensity in each channel from my z-stacks for comparison.
Unfortunately, there are some samples with very weak EdU labeling, where the exposure time is very long (~6-8 seconds vs ~175ms for dapi), so my intensity appears somewhat high everywhere (not just in nuclei), due to weak auto-fluorescence.
Is there any way for me to control for this difference in exposure time or fluorescence localization?
Thank you!
Try to increase the digital gain of the camera in the "EdU channel" and to subtract the background from the channel histogram using min/max or best fit. Strange the EdU usually gives strong signal. Think also ways to improve your staining protocol?
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