Leukocyte Alkaline Phosphatase

Created by Kathleen Patrick, Modified on Fri, 8 Nov at 12:54 PM by Kathleen Patrick

EMS26913

Components

Buffered Acetone Fixative

Naphthol AS-BI Phosphate

Dimethyl Formamide

Fast Red Violet Salt LB

Propanediol Buffer

Haematoxylin Counter Stain

Sample

Capillary Blood Or Heparinized Venous Blood Films

Fixative

Buffered Acetone Fixative

Procedure

Fix blood films in Buffered Acetone Fixative for 30 seconds.
Wash Blood films in running tap water for 30-60 seconds. Air dry or blot.
Stain films for exactly 10 minutes in staining Solution. *To prepare staining solution: Dissolve 5mg of Naphthol AS-BI Phosphate in 0.2 to 0.3ml of Dimethyl Formamide. Add 60m of Propanediol Buffer. Add 40mg Fast Red Violet Salt LB or Fast Violet B Salt. Filter and use at once.
Wash blood films in running tap water for 30 to 60 seconds.
Counterstain in Haematoxylin Solution filtered just prior to use, for 5 - 8 minutes.
Rinse in running tap water 1 to 2 minutes. Air dry or blot carefully.
Examine microscopically under an oil immersion lens.

Stain Results

Intensity of precipitated dye in the cytoplasm of neutrophilic leukocytes varies. One hundred are scored from 0 to 4+ based on the intensity of the precipitated dye.

Scoring Criteria

CELL RATING	AMOUNT **%	Precipitated azo dye In cytoplasm Size	INTENSITY	BACKGROUND OF CYTOPLASM
1+	<50	Small	Faint to moderate	Colourless to very pale pink
2+	50-80	Small to Med.	Moderate to strong	Colourless to pale pink
3+	80-100	Med. To Large	Strong	Colourless to pink
4+	100	Med. To Large	Brilliant	Not visible

* When there is no precipitate azo dye in the cytoplasm and the background is not stained, the cells are read as zero.
** Percentage of volume of cytoplasm occupied by azo dye precipitate.

Normals

Adults - Leukocyte Alkaline Phosphatase Score Of 13 To 130.

References

Williams, W.J., Beutler, El, Erslev, A.J. and Rundles R.W., Hematolog y, 2 nd ed., McGraw-hill , NY , C. 1977, p1628-1629.

Kaplow, L.S., AM.J., Clin. Patbol ., 39:439, 1963.

Kaplow, L.S. and Burstone, M.S., Nature , 200:690, 1963.