Flow Cytometric Assessment of Neutrophil Oxidative Metabolism in Chronic Granulomatous Disease on Small Quantities of Whole Blood: Heterogeneity in Female Patients

A rapid and sensitive flow cytometric assay is presented for the quantitative estimation of the oxidative metabolic activity of individual polymorphonuclear leukocytes (PMN) on less than 100 td of whole blood. This procedure is a simplified version using whole blood of the method of Bass et al (J. Immunol. 130:1910, 1983) that estimated the metabolic burst activity of phorbol myristate acetate (PMA)-stimulated individual PMN as the intracellular generation of a fluorescence product by a flow cytometric assay. With this method, almost all the PMN from normal subjects responded to PMA as a single cell population generating bright intracellular fluorescence. PMN from a boy with chronic granulomatous disease (CGD), could not respond to PMA with any increase of their fluorescence intensity. His mother had two distinct PMN populations one functionally normal and the other defective, indicating a random lyonization in the carrier mother and the X-linked recessive mode of inheritance. In two female patients with CGD from unrelated families, their PMN responded to PMA, as a whole, with a minimal increase in the fluorescence intensity, but the metabolic defects in their PMN were not so complete as seen in a classical X-linked CGD boy. But, PMN from two female sibling patients from the other family responded to PMA as a single uniform cell population with a weak but definite fluorescence intensity. However, the genetic background of these female patients with CGD remains unclear, since PMN dysfunction could not be identified in their mothers with this method.