Hello everyone,
I'm currently working on immunofluorescence staining experiments, and I've noticed a consistent issue. Regardless of the primary antibody I'm using, I always observe higher background staining in the epidermis compared to the dermis.
I'm careful with my blocking steps and have tried different blocking agents, but the problem persists. Has anyone else experienced similar issues, or does anyone have insights into why the epidermis might show higher background staining? Any suggestions on how to reduce this would be greatly appreciated.
Thank you in advance for your help!
Hi, Zahraa,
I am wondering which type of skin you work on. Is it human or mouse skin? I have done a lot of skin including pig, human and mouse skin for immunofluorescent staining. The skin from pig and mouse usually have no background in dermis and epidermis, but I usually see a lot background in human skin. I don't know the reason but it seems relative to blood remain in the skin, thickness of sections and hard to de-wax in human skin. you may try to remove the blood before fixation to reduce the background. in mouse skin, if you see a lot background in epidermis, it usually come from the secondary antibody. you may try to change the secondary antibody.
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