Cytochemistry of Leukocyte Alkaline Phosphatase: Use of Complex Naphthol AS Phosphates in Azo Dye-Coupling Technics

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Abstract
The substituted naphthol AS- BI phosphate coupled with Red Violet Salt LB, buffered to pH 9.7 with 0.05 M 2-amino-2-methyl-l,3-propanediol yields a particulate chromogenic red azo dye within the cytoplasm of human blood neutrophiles. "Scoring" is simplified by the ease of detection of faintly stained cells. The use of other naphthol phosphates and other diazonium salts gave less satisfactory results. A frozen stock substrate solution was stable for over six months. A greater differential was noted between the average "scores" of healthy adults and patients with chronic myelogenous leukemia as compared to results obtained with the unsubstituted naphthols. The concentration of naphthol and diazonium salt could be varied within wide limits without effecting the appearance of the azo dye. The effects of fixation, storage, incubation time, temperature and pH on enzyme activity are described. The method is rapid, simple, inexpensive and reproducible.