[Histonet] (kinda) silly questions about IHC

[Anna Elisse Beaudin]

Hello,

   I have a question about IHC.  I am trying to do multiple labeling on
sagittal brain sections that were fixed by perfusion.  I cut these
sections on a cryostat, and I am having a lot of trouble deciding
whether to do IHC on mounted sections vs. free-floating.  When I try to
mount fixed sections directly while sectioning, I get air bubbles
caught between the section and the slide.. the sections just don't seem
to want to lie smoothly on the slide.  Alternatively, when I stain
free-floating sections, these long sagittal sections curl up and as
such I get extremely uneven (and ugly) staining.  My actual (silly)
question is whether it is possible to collect sections into solution
(free-floating), mount them, let them dry overnight, and then do IHC on
them.  I'm torn because I need to let the section dry a little on the
slide so that they stick, but at the same time I think it might be bad
for them to dry out?  As you can see, I am very confused, and would
really appreciate anyone's advice.  Thanks in advance for your help!

Best,
Anna Beaudin
Division of Nutritional Sciences
Cornell University