Cytochemical flow analysis of intracellular G6 PD and aggregate analysis of mosaic G6 PD expression

Abstract Background Medicines that exert oxidative pressure on red blood cells ( RBC ) can cause severe hemolysis in patients with glucose‐6‐phosphate dehydrogenase (G6 PD ) deficiency. Due to X‐chromosome inactivation, females heterozygous for G6 PD with 1 allele encoding a G6 PD ‐deficient protein and the other a normal protein produce 2 RBC populations each expressing exclusively 1 allele. The G6 PD mosaic is not captured with routine G6 PD tests. Methods An open‐source software tool for G6 PD cytofluorometric data interpretation is described. The tool interprets data in terms of % bright RBC , or cells with normal G6 PD activity in specimens collected from 2 geographically and ethnically distinct populations, an African American cohort ( USA ) and a Karen and Burman ethnic cohort (Thailand) comprising 242 specimens including 89 heterozygous females. Results The tool allowed comparison of data across 2 laboratories and both populations. Hemizygous normal or deficient males and homozygous normal or deficient females cluster at narrow % bright cells with mean values of 96%, or 6% (males) and 97%, or 2% (females), respectively. Heterozygous females show a distribution of 10‐85% bright cells and a mean of 50%. The distributions are associated with the severity of the G6 PD mutation. Conclusions Consistent cytofluorometric G6 PD analysis facilitates interlaboratory comparison of cellular G6 PD profiles and contributes to understanding primaquine‐associated hemolytic risk.