We also do this. A couple of additions/suggestions/hints:

First, the potassium permanganate/sulfuric acid solution
tends to pull the tissue off slides. Place the tissue on
subbed slides (plus, poly-L-lysine, silane, etc. - the
stronger adhesive, the better) and dry the slide/tissue for
several hours before staining. Then, be VERY GENTLE
with the washes in water.

Second, to work best, one needs both AA amyloid
control and AL amyloid control.

So there are 6 slides - 1 set (AA control, AL  control
and patient slide) to go through the pretreatment, and
1 set to NOT go through treatment (stay in water
until hematoxylin step).

We have to prove that the Congo red will stain both
AA and AL amyloid, and that the pretreatment will
stop the AA amyloid from staining, but that the AL
amyloid will continue to stain. Then compare the
patient's tissue to the two controls.

The problem is, AA amyloid (secondary amyloidosis)
controls can be difficult to find.

Third, AL amyloid will continue to stain after pre-treatment,
but the staining may be REDUCED in staining, anywhere
from slightly reduced staining to greatly reduced staining.
However, the AA amyloid will not stain at all. So it can get
tricky - is the patient's tissue totally not staining, or is it
staining but greatly reduced staining? Sometimes it's
almost a matter of interpretation.

Has immuno been considered? Mouse anti-Human
Amyloid A Component will stain AA amyloid.

Peggy A. Wenk, HTL(ASCP)SLS
William Beaumont Hospital
Royal Oak, MI 48073

----- Original Message -----
From: "Andrew Kennedy" 
To: 
Sent: Wednesday, January 23, 2002 4:43 PM
Subject: Re: AL vs AA amyloid


> Hi Jill,
>
> We use an acidified permanganate solution as pretreatment for the Congo
Red
> stain to distinguish secondary amyloid (composed predominantly of protein
> AA) from other forms.
>
> Controls:
> 2 control sections and 2 sections from test case are used - one control
and
> one test slide is kept in water until step 5.
>
> Solutions:
> 1.     Acidified permanganate solution:
>             0.25% sulphuric acid - 25 mL
>             5% Potassium permanganate - 25 mL
> 2.    5% Oxalic Acid
> 3.    Congo Red in 50% EtOH
> 4.    0.2% KOH in 80% EtOH
>
> Method:
> 1.    Take sections to water
> 2.    Place in acidified permanganate solution 2.5 to 3 mins
> 3.    Wash in water
> 4.    Clear in Oxalic acid
> 5.    Wash well in water
> 6.    Stain in Haematoxylin, differentiate and blue
> 7.    Wash well in water
> 8.    Stain in Congo Red - 15 min
> 9.    Differentiate in KOH - 10-15 sec
> 10.  Rinse in water, dehydrate, clear and mount
>
> Results:
> The secondary amyloid loses Congo Red affinity and polarisation
> characteristics...
>
> The reference quoted in our manual for this is: Wright et al, (1977). Lab
> Invest 36(3); 274-281.
>
> Hope this helps you out....
>
> Andrew Kennedy
>
> Senior Science Officer in charge - Histopathology.
> Department of Anatomical Pathology
> Concord Repatriation General Hospital
> Hospital Road
> Concord, NSW, Australia
> 2139
> Phone: +612 9767 6115
> Fax: +612 9767 8427
>
> "Noli illegitimi carborundum"
>
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