Hello everyone,

We have been staining our single cell slides with DAPI and mount the slides with vector mounting media that contains anti-fade reagents. Immediately after staining, DAPI and all the fluorochromes are clearly visible. However, after merely 2-3 days, I usually find the DAPI staining starts to turn pink when the other fluorochromes remain strong. In fact, it seems that the DAPI blue is turning up around the cells but not inside the cells. So I ended up with faint pink outlining the nucleus and blue all around, making it very difficult to take photos. I am not sure if in this case DAPI is considered "faded" since I can still see blue stain even though they are outside of the cells. I wonder whether it is the fixation that is causing the problem? This problem doesn't seem to be as serious in parafilm sections. Please help. Thanks!!

Why do you have to check the immunostaining after 2-3 days. We usually check the fluorecense right away or at most after 1 overnight.