My first question is in what region of skin is the staining poor? The
fibroblasts of the dermis should stain really well with H&E = as
should the cells of the stratum germinativum. From there out nucle ar
staining gets worse and worse. I attribute it to two things, and
i= f anyone else would add I would love more insight into this
problem: = First is keratinization. Since cells are no longer
dividing and are g= radually dying and losing nuclei as they make
their way out to the stratum = corneum, nuclear staining is going to
be sparse as a result of the process = itself. Also, highly
keratinized epithelia tend to be problematic for= many embedding and
staining techniques where tissue has to be dehydrated a= nd
rehydrated. I have always had trouble sectioning highly keratinize= d
skin because of improper infiltration. Second is the organization o f
chromatin in epidermal cells in general, which is seemingly very
differen= t than other epithelial cells (again, the stratum
germinativum often does s= how typical nuclear staining).
With all that being said, I hav= e tried several things to remedy
problems associated with H&E staining = of skin with mild success.
Sometimes it might be as simple as adding = a few drops of glacial
acetic acid to your hematoxylin to lower the pH.&nbs= p; When this
doesn't work, I typically will use iron alum or ferric chlorid= e (I
believe 2-4% iron alum has worked relatively well for me when used)
as= a mordant prior to staining with Delafields or Ehrlich's
hematoxylin. = ; Destain with 2-4% iron alum or ferric chloride
because anything holding t= he alum when you stain picks up
hematoxylin and will need to be differentia= ted before
counterstaining. I have also had mild success with iron ga= llein
elastin stain, but this is really more of a stain for elastin that ju st
happens to stain nuclei blue-black and gives good
differentiation. = Gallein is also not as easy to find as Hematoxylin.
As I said= , if anyone has information on either the chemistry
behind this or maybe ev= en on the organization of chromatin in
epidermal cells, the histo community= is all ears.
-Matt
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