again blueing tissue sections

From:Heike Grabsch <> (by way of histonet)

First of all, I was amazed to get so many answers instantly via the
histonet. I posted the message in the evening after coming home
frustrated and already had a couple of answers before I went back to
work next morning.

There were a couple of suggestions made I would like to comment, because
my problem is not solved after todays experiments:

- yes, I made my own Mayer's and filtered it after all chemicals were
solved, however I do not filter it every day

- I have already seen precipitates coming directly from haemalaun (or is
it called haematoxylin? please forgive me for my ignorance), in my
experience they have some sort of right angles and sharp borders and
often are a little bit yellowish in the edge

- may be,  I should describe my procedure again in more detail (like I
did it today): so after IHC staining, I use AEC one step solution
(BioGenex) as chromogen, rinse in dist. water, 90 sec Mayer's, first
rinse in dist. water, then extensive rinsing with running cold (!) tap
water, then alternatively 30 sec 0,5% Lithium carbonate or 0,25% ammonium
hydroxide as suggested by Frieda this morning, thereafter extensive
rinsing in running cold tap for may be 30 minutes, BECAUSE on both
versions I had again these branched and spinous, deep blue precipitates
and could not get rid of them. And of cause they are not all over the
tissue section but mainly on top of the cells of interest! And you only
see them on the tissue, no precipitate on the slide itself.

As nobody else seem to have this problem, I must make a major mistake.
So, from today's experiment I conclude it is probably not the lithium
carbonate. So, may be it is something in our tap water? I will make my
next trial tomorrow using only dist. water for rinsing. Let's hope for
the best.

any more ideas?

Heike Grabsch, Germany

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