I do IF every week with different antibodies on transverse E12.5-E14.5 mouse embryo sections. One thing I commonly noticed is that the signal in 90% cases is misleading. For example, PDGFRa signal should be in the entire dermis but I see it either in the left or right half of the dermis and not the other half. Am I doing something wrong?
Hi Venkata,
If you think that all your sections were treated exactly the same (and the whole section) my suggestion would be to make sure youf UV light source is aligned correctly. It could be that is not centered and that would explain why one side does not have signal. The easiest way of testing this is rotating the sample as you look at the image, if misaligned you should see the fluorescence appear/disappear as the sample gets in the path of the UV light.
Hope this helps,
Ezequiel
Check your microscope shuttle which work properly. I had similar problem and it was the microscope.
Another case I have come across was that the objective turret was incorrectly positioned.
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