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 Heparinised 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.