A RAPID SINGLE-STEP SCREENING METHOD FOR GLUCOSE-6-PHOSPHATE DEHYDROGENASE DEFICIENCY IN FIELD APPLICATIONS

Abstract

The single-step screening method (SSS) is a qualitative rapid screening test for glucose-6-phosphate dehydrogenase (G6PD) deficiency based on blue formazan formation on anion-exchanger.The reaction mixture contains equal volumes of anion exchanger, substrate mixture, coloring mixture (MTT-PMS mix) and distilled water. We assessed the stability of the reaction mixture and evaluated its reliability with two anion exchangers, DEAE-Sephadex A50™ and DEAE-Sephacel™, for applications in tests under field conditions. The reaction mixture was sufficiently stable under conditions of incubation at 70°C for 6 hours or vigorous shaking for 24 hours at room temperature. The reaction mixture could be kept at 30-35°C for 14 days under indoor conditions without shielding if it contained no MTT-PMS mix. The coloring was detectable even in diluted blood with hemoglobin concentration as low as 1.6 g/dl. Under laboratory conditions, the proportion of the samples with 10% of the normal level of activity that were diagnosed as 'low activity' was higher with DEAE-Sephacel (92%) than with DEAE-Sephadex A-50 (81%) (p=0.023). The proportion of the samples with normal activity that were diagnosed as 'normal' was 98% with DEAE-Sephacel and 100% with DEAE-Sephadex A-50. In field samples obtained in Myanmar and Indonesia, the sensitivity was lower (P=0.03 using DEAE-Sephadex and p< 0.001using DEAE-Sephacel) when we used the blue formazan spot test (BFST) as the standard. Twenty-three of 27 G6PD-deficient individuals subjected to genetic analysis were found to have mutations. All individuals who had concordant results between the SSS and the blue formazan spot test (BFST) carried molecular mutations. One case of G6PD mutation was detected among four cases diagnosed as G6PD-deficient by SSS with DEAE-Sephacel™, but diagnosed as 'normal' by BFST. The costs of one test with the DEAE-Sephadex A-50™ and the DEAE-Sephacel™ system were 0.15 US dollar and 0.30 US dollar, respectively. (297 words)