You are using less acid fuchsine than is usual in
Van Gieson's stain. Usually the mixture contains 0.1%
of the dye in saturated picric acid. Starting with
a 1% solution you'd therefore need to add 5 ml to
45 ml of saturated picric acid. An optional additive
is 0.5% (0.25 ml) of conc. HCl. Add this if the red 
colour is spilling over into cytoplasm. It should be
only in collagen fibres. The thinnest fibres do not
stain with Van Gieson's method. It is easy to lose
the collagen stain. Rinse well in slightly acidified 
water (never in tap water) to remove unbound dye,
and dehydrate in 3 changes of 100% alcohol (not
graded alcohols). Too long in alcohol can remove
the picric acid component of the stain.

Once you've got it working you can do great batches
of slides with Van Gieson, and the staining solution
seems to last for ever. It's an excellent oversight
stain - much better than H&E and easier to do well.

-- 
-------------------------
John A. Kiernan
Department of Anatomy and Cell Biology
The University of Western Ontario
London,   Canada   N6A 5C1
   kiernan@uwo.ca
   http://publish.uwo.ca/~jkiernan/
_________________________________________________
NIDAL E MUVARAK wrote:
> 
> Hello Histonetters. I had a question regarding collagen staining in VVG stain. I'm following a protocol where the slides are incubated for 30 minutes in Verhoeff's hematoxylin, differentiated in 2% ferric chloride, rinsed in hypo solution to remove iodine, then counterstained in Van Gieson's solution for 5 minutes. The results should show black elastic fibers and nuclei and red collage on a yellow background (other tissue elements). I do get those results, however the collagen stains a very faint red. The Van Gieson solution is made with 1ml 1% Acid fuchsin (1gm acid fuchsin in 100ml distilled water) and 45ml saturated picric acid, and allowed to stand overnight before staining. Now my question, is this the best way to make Van Gieson's solution, and if it is, should I just counterstain with it for longer than 5 minutes to get a brighter red? I know these might be trivial questions, but I've never done this stain but once recently and I don't have the time to troubleshoot !
 sin
> ce my PI wants the results for a paper ASAP. Thank you for your time.
> 
> Nidal E Muvarak
> Associate Research Specialist
> Department of Biomedical Engineering
> University of Wisconsin-Madison
> 1550 Engineering Dr.; Rm. 2158
> Madison, WI 53706-1609
> Lab: (608) 265-8921; Office: (608) 265-4205;
> Home: (608) 256-7934; Cell: (608) 332-6068