Kinetic analysis in single, intact cells by microspectrophotometry: evidence for two populations of erythrocytes in an individual heterozygous for glucose-6-phosphate dehydrogenase deficiency.

A microspectrophotometric technique was used to measure the kinetics of methemoglobin reduction in intact, unaltered human erythrocytes. Reduction was catalyzed by endogenous NADPH-methemoglobin reductase in the presence of Nile Blue. The technique was applied to the study of erythrocytes from a female donor with decreased glucose-6-phosphate dehydrogenase (G-6-PD) activity. The individual was shown to be heterozygous for deficiency of G-6-PD. The kinetic study revealed two distinct populations of erythrocytes that were nearly equal in number. One cell population showed reduction rates between 0 and 25% of normal, whereas the second cell population displayed rates within the range seen for normal cells. Single-cell indices of cell size, cell hemoglobin content, and ratio of cell hemoglobin to cell size did not correlate with single-cell reduction rates and were not significantly different between the two populations. These results provide quantitative support for the X-inactivation hypothesis in G-6-PD deficiency.