SEMINAR ON PAS

PRESENTED BY :-Ms ANJANA B.Sc .MLT 3rd yr MODERATED BY:- Dr AMAR RANJAN

PERIODIC ACID SCHIFFS REACTION :- [ PAS ]

Detects

intracellular glycogen and neutral mucopolysaccharides in variable quantities in most hematopoietic

found

cells.
Staining

is seen in blast of acute lymphoblastic leukemia.

Diffuse

cytoplasmic positivity in erythroleukemia. useful in demonstrating the abnormal glucocerebrosidase accumulation in GAUCHERS disease.

Very

PRINCIPLE

PAS reaction depends upon the liberation of carbohydrate radicals from the combination with protein and their oxidation to aldehydes by the schiff s reagent.

RESULTS AND CONFIRMATION

Positive reaction denotes the glycogen presence Reaction can be confirmed by demonstrating that the positive reaction disappears on treatment with diastase or saliva before staining

other PAS positive material is changed by diastase digestion.

REACTION

REAGENTS USED AND PREPARATION

PERIODIC ACID SOLUTION DISTILLED WATER SHIFFS REAGENT COUNTER STAIN HEMATOXYLIN

1g 100 ml

PREPARATION OF SCHIFFS
Dissolve 1 g basic fuchsin in 200 ml of boiling, removing the flask of water from the bunsen just before adding the Basic fuchsin. Allow the sol. to cool to 50c & add 2 g potassium metabisulphite with mixing. Allow to cool then add 2 ml conc. HCL, mix & add 2g activated charcoal & leave overnight in the dark at room temperature . Filter & store at 4c in amber coloured bottle.

 FIX THE SMEAR IN METHANOL TREAT WITH PERIODIC ACID FOR 10 MINUTES

WASH WELL IN RUNNING WATER

COVER WITH SCHIFFS REAGENT FOR AN HOUR

COUNTER STAIN WITH HAEMATOXYLIN STAIN WASH AND RINSE IN ALCOHOL

RINSE IN DISTILLED WATER

CLEAR IN XYLENE MOUNT IN DPX

GIANT MULTINUCLEATE CELLS SHOWING GRANULAR POSITIVITY

INTERPRETATION

Normally all blood cells are (+ve) but Erythroblasts (-ve)

Diffused stain pattern (Granulocytes) - Granular stain (lymphocytes and monocytes) - Plts deeply stained - nRBCs (-ve) stain

In diseases: In CML Lymphosarcoma Hodgkins disease nRBCs in M6 Thallasemia

Positive PAS stain in ALL

PAS positivity in M6. Not the intense staining of the large abnormal erythroblast.

CLINICAL SIGNIFICANCE
POSITIVITY

IS SEEN IN :B CELLS :- Strongly +ve T Cells :- Weakly +ve

Deep

diffuse staining in erythroleukemia & in thallasemia.

Lesser

a)
b)

c)
d)

e)

degree of staining may be seen in Iron deficiency anaemia Myelosclerosis Cord blood erythroblasts Sideroblastic anaemia In various type of leukemia & hemolytic anaemia.

Reaction

is +ve in a) Pernicious anaemia b) Aplastic anaemia c) Lead poisoning d) Polycythemia Vera

NOTE

a)
b) c) d)

Reaction is best carried out on Fresh blood Bone marrow films Old methanol fixed films Films stained by Romanowsky dyes, months or before can be quite satisfactory.

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