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Electron Microscopy Sciences

Technical Data Sheets

Leukocyte Alkaline Phosphatase

EMS Catalog #26913

Sample

Capillary Blood Or Heparinized Venous Blood Films

Fixative

Buffered Acetone Fixative

Procedure

  1. Fix blood films in Buffered Acetone Fixative for 30 seconds.
  2. Wash Blood films in running tap water for 30-60 seconds. Air dry or blot.
  3. 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.
  4. Wash blood films in running tap water for 30 to 60 seconds.
  5. Counterstain in Hematoxylin Solution filtered just prior to use, for 5 - 8 minutes.
  6. Rinse in running tap water 1 to 2 minutes. Air dry or blot carefully.
  7. Examine microscopically under an oil immersion lens.

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 **% Ppt'd azo dye In cytoplasm Size INTENSITY BACKGROUND OF CYTOPLASM
1+ <50 Small Faint to moderate Colorless to very pale pink
2+ 50-80 Small to Med. Moderate to strong Colorless to pale pink
3+ 80-100 Med. To Large Strong Colorless 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.

Online Ordering

Buffered Acetone Fixative, Naphthol AS-BI Phosphate, Dimethyl Formamide, Fast Red Violet Salt LB, or Fast Violet B Salts, Propanediol Buffer and Hematoxylin Counter Stain for Leukocyte Alkaline Phosphatase are available online from the EMS Catalog. For ordering or product information, click here.