711 DIAGNOSING THE CARRIER STATE IN CHRONIC GRANULOMATOUS DISEASE (CGD)
1978
Although laboratory diagnosis of CGD presents no difficulty, present techniques are insufficiently sensitive to detect those CGD heterozygotes in whom the majority of cells are functionally normal. We modified the test proposed by Preisig and Hitzig (Eur. J. Clin. Invest. 1:409, 1971) to improve its sensitivity and accuracy. Peripheral blood leukocytes are exposed to heat killed C. albicans in the presence of 0.05% nitroblue tetrazolium (NBT) and 8% Group AB serum for 15 min. at 37C. Next, an equal volume of 0.2% eosin Y in phosphate buffered saline is added for 15 additional min. The cells are centrifuged, resuspended in a saline solution and examined microscopically. Noningested yeasts whether extracellular or adherent to the cell's surface, are stained red. All intracellular yeasts within normal neutrophils or monocytes are stained blue, by virtue of their content of reduced NBT. Yeast cells within neutrophils or monocytes from patients with CGD lack reduced NBT and appear white in color. Heterozygotes have two populations of leukocytes, one (normal) that contains blue yeast cells and the other (CGD) that contains white ones. In two heterozygous females we studied, only 11.5% and 26% of the neutrophils displayed the CGD phenotype. With the use of eosin Y to stain noningested Candida cells our method is sufficiently sensitive to detect heterozygotes with as few as 1-2% defective cells. We speculate that some females with clinical CGD may prove to be such heterozygotes.
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