microwave PAS stain

From:Steven Slap

Hi HistoNetters

Rita Angel asked about microwave PAS staining.  Without knowing what kind of
microwave Rita has, it is hard to give advice, but the following protocol,
developed by Joyce Moore, one of the microwave staining pioneers, is very
reliable in my hands:

Periodic Acid Schiff Reaction
Fixation: Formalin
Technique: Cut paraffin sections at 4 microns
Control: Tissue containing PAS positive material
Solutions: 
0.5% Periodic Acid Solution

Periodic acid crystals 0.5 g

Distilled water 100.0 ml


Schiff's Reagent

Pour approximately 40 ml of Schiff solution in coplin jar. Store in
refrigerator. Discard when solution becomes pink.

0.5% Diastase solution light green

If staining for glycogen, use liver with glycogen in it for a control.


Staining Procedure:
1.    Deparaffinize and hydrate sections to distilled water with the aid of
the automatic stainer.
2.    Rinse in distilled water.
3.    If PAS reaction with digestion is desired, place sections in coplin
jar in 0.5% diastase for 45 seconds in microwave. Parallel slides should be
placed in distilled water in coplin jar and microwaved for 45 seconds. Rinse
slides in running tap water. Rinse in distilled water.
4.    Periodic acid solution for 10 seconds in microwave.
5.    Rinse in distilled water, 2 changes.
6.    Place in Schiff's reagent for and microwave for 30 seconds (Note: in a
laboratory microwave, use 100% power at 30 C for 30 seconds). In a
laboratory microwave, stir solution while heating with an air bubble
agitator. Otherwise, remove from microwave and agitate the solution by hand
to equilibrate the solution temperature. This will allow for more even
staining. Let stand in stirred solution for 30 seconds to 5 minutes after
removing from microwave, until sections have assumed a pinkish color. Check
with microscope for correct staining.
7.    Wash in running water for 5 minutes.
8.    Attach to automatic stainer. Stain in Hematoxylin solution for 15
seconds. 
9.    Rinse in water.
10.    Differentiate in acid alcohol.
11.    Rinse in water.
12.    Blue in running tap water.
13.    Rinse in water.
14.    Dehydrate, clear. Mount with xylene soluble media.

Results:
Glycogen - rose to purplish red
Mucopolysaccharides, fungi - rose to purplish red
Background - blue
Adapted for the Jefferson Regional Medical Center Histo-Path Laboratory

best regards,
Steven Slap

**********************************************
Marketing Manager/Microwave Product Specialist
Hacker Instruments & Industries, Inc.
http://www.hackerinstruments.com
**********************************************
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