Image: The qualitative point-of-care test CareStart G6PD RDT is a visual screening test that identifies G6PD deficient patients using whole blood sample. Panel A, no color change for sample with deficient G6PD enzymatic activity; Panel B, distinct purple color for sample with normal G6PD enzymatic activity (Photo courtesy of Access Bio).
Glucose-6-phosphate dehydrogenase deficiency (G6PDd) is the most common enzymopathy with at least 400 million individuals affected worldwide. More than 185 clinically relevant variants of G6PDd have been reported with a spectrum of associated enzyme deficiencies.
The presence of G6PDd in patients with Plasmodium vivax infection results in significant challenges to achieving radical cure. The 8-aminoquinolone compounds are the only class of anti-malarial drugs currently available to eliminate the dormant hypnozoites liver stages of P. vivax are and these cause hemolysis in G6PDd individuals.
Scientists at the Charles Darwin University (Sydney, Australia) and their colleagues reviewed the methods for the field evaluation of quantitative G6PD diagnostics. G6PD enzyme activity and its degradation is temperature dependent, hence whole EDTA blood should not be stored at room temperature. Freezing samples and maintaining a respective cold chain in a field setting is demanding and often impossible. Whole blood samples can be stored at 4–8 °C for later analysis; however the duration at which samples can be stored and reliably assayed at a later time point is unknown. Field evaluation is based on the comparison of paired samples, including one sample tested by the new assay at point of care and one sample tested by the gold-standard reference method, UV spectrophotometry in an established laboratory.
In one study the G6PD activity of paired samples collected from venous and capillary sampling were similar when measured by spectrophotometry, the medians were 7.51 and 7.61 U/gHb, respectively. Similarly, an evaluation of qualitative tests by fluorescent spot test (FST) and Carestart G6PD RDT showed 100% concordance between paired capillary and venous samples. ACCESS BIO has recently developed a Biosensor, a quantitative handheld device that measures G6PD activity based on the electrochemical properties of G6PD in blood samples.
The WST8/1-methoxy PMS test is a robust and cost effective alternative to estimate G6PD activity that can be performed on dried blood spots and whole blood samples. The WST8/1 test is based on the formation of an orange formazan dye generated by G6PD enzyme activity and facilitated by the 1-methoxy PMS as an electron carrier. The assay has been adapted for quantitative batch testing in a 96 well microplate and can be read by an enzyme-linked immunosorbent assay (ELISA) reader.
The authors concluded that their review presented a number of recommendations to facilitate standardized evaluation of quantitative G6PD activity test assays. Standardized evaluation studies are necessary to provide a comparable basis for subsequent pooled analyses. As novel quantitative G6PD diagnostics become available their potential to provide robust quantitative G6PD results suitable for field deployment needs to be assessed. The most recent generation of instruments contain an integrated temperature correction factor and hemoglobin measurement and G6PD activity is displayed normalized as U/gHb. The study was published on September 11, 2017, in the Malaria Journal.
Charles Darwin University