Dear BioForum users,
I have recently started performing immunofluorescence on fixed mammalian cells.
Although the steps are pretty straightforward, the coverslips carrying the cells would always tend to dry out between the different incubations.
As a result the antibody solutions won't spread well and evenly on the cell layer. I am afraid that this problem generates artificial differences in the staining intensity besides the high background.
I always try to leave some small amount of fluid (eg PBS) from the previous steps on the coverslips but on the other hand I am afraid that this might overdilute my antibody and distort the initial concentration.
Could anyone please suggest ways to avoid drying of the cells between the different IF steps? Any help would be really appreciated.