Hi Joseph and all,

By your question, I'm not sure if you are having problems with the stain
or with removing the stain by hyaluronidase digestion. If it's the
digestion step that you are having problems with, I had good luck with
Citrate/phoshate buffer, 0.1M at pH 5.8 and digestion in the oven at 37o
C.
 My process went overnight, as I was digesting through the celloidin
embedding media, but it worked well.  I know the oven is important because
our oven was turned off in the middle of one run and the tissue did not
digest.

I hope this helps.

Karen Pawlowski
Sr. Research Assoc./UT Southwestern Med. Ctr.;
PhD Candidate/ UT Dallas, Dallas, TX


On Fri, 26 Feb 1999 FRUGE.JOSEPH@TUCSON.VA.GOV wrote:

> We have consistently had difficulty with the Alcian Blue with and without
> Hyaluronidase Stain.  We are using a section of cartilege as our positive
> control. We use a Phosphate buffer solution at pH 5.6 with testicular
> hyaluronidase purchased from Sigma Chemical.  We use 25 mg of
> hyaluronidase to 50 cc of buffer. We are using a 3% solution of Alcian
> Blue ph 2.5.  Any help out there????
>
>