Table 1.

Available tests for determination of G6PD deficiency and their use in field settings

Test	Characteristics	Shortcomings for field and mass-screening
DNA sequence analysis of the G6PD gene.	Extremely reliable. Primers are used to check whether the G6PD gene contains a mutation.	Requires training, and equipment. Genotype does not correlate with enzyme function and the risk of haemolysis. Female heterozygous have unpredictable phenotype due to X chromosome lyonization. Only one mutation can be analysed with one primer (>160 mutations exist).
Brilliant cresyl blue decolouration test	Involves the action of G6PD and NADPH diaphorase. A deficiency of either one of these enzymes on RBCs would result in the brilliant cresyl blue remaining unchanged in the test.	Laborious processes; requires technical skill, and has low sensitivity.
Methaemoglobin reduction test	Based on the oxidation of Hb to MetHb by sodium nitrate and the subsequent enzymatic reconversion to Hb in the presence of methylene blue.	Laborious, qualitative and low sensitivity. Does not enable identification of heterozygous deficient females.
Formazan ring method	Uses the principle of the MTT-Linked spot test. When G6PD is present at normal levels, MTT is reduced to a purple insoluble formazan derivative, and results in a specific diameter of discolouration.	Prone to misdiagnosis.Ring thickness may be affected by exogenous factors.
Sephadex gel MTT-PMS method	Mostly used in Asia, and predecessor in concept, of the WST8/1-methoxy PMS test.	Reacts with haemoglobin; is light sensitive and water insoluble. It is of a qualitative nature.
Fluorescent spot test (FST)	ICSH-recommended method.	Its cut-off value for G6PD deficiency determination is only 10-20% of the normal G6PD activity, which excludes patients with moderate enzyme deficiency and increases the risk of false-normal diagnosis.
BinaxNOW® rapid test	Rapid test format: Overcomes issues of technical skill, sophisticated equipment and reliability.	It is highly dependent on temperature-sensitive kinetic enzymatic reactions. This limits its use to areas with temperatures between 18 and 25C. Potential cost.
CareStart™ test	RDT format. Qualitative chromatographic test, based in the reduction of colourless nitro blue tetrazolium dye to dark colour formazan. Long-term temperature stability.	Potential cost.
R&D® enzymatic test (reference)	Both depend on the conversion of NADP + to NADPH by G6PD. NADPH converts colourless tetrazolium salt into a coloured formazan, while NADP + does not.	Enzymatic gold standard. Requires various temperature-dependent incubations.
WST8/1-methoxy PMS test (test under validation)		Evaluated in this work. Advantages: no reaction with haemoglobin, lower light sensitivity.