G6PD Assay Kit - WSTG256 Product Manual

Glucose-6-phosphate dehydrogenase (G6PD) deficiency, the most common enzymopathy, causes acute hemolytic anemia. Therefore the measurement of G6PD activity is important, particularly in malaria-endemic areas where primaquine is widely used as an anti-malaria drug in the treatment of malaria. G6PD Assay Kit-WST utilizes a water-soluble tetrazolium salt, WST-8, that also produces water-soluble formazan with an intense orange color at 460 nm. WST-8 does not react with hemoglobin, allowing a very simple and rapid assay for screening of G6PD activity.

20 µl and 1 ml micropipettes and tips 1.5 ml microcentrifugation tubes water (commercially available, fresh drinking water) 1 mol/l HCl.

1. Add sequentially 760 µl of water, 20 µl of Substrate Mixture and 20 µl of Dye Mixture into 1.5 ml tubes.
2. Add 5 µl of sample bloods into 1.5 ml tubes.
3. Mix them vigorously for 5 sec and incubate at 25~37°C for 20~30 min.
4. Stop the reaction by adding 10 µl of 1 mol/l HCl and compare the developed color with those of positive and negative controls.

Orange color development (HCl is not added)

1. Store the Kit at -20°C. This kit is stable for 6 months at -20°C, for one month at 4°C,and for ten days at 37°C .
2. After thawing Substrate Mixture and Dye Mixture, it is recommended to mix them with 76 ml of water (total 80 ml) and dispense 800 µl of the mixture to 1.5 ml tubes as described in the assay procedure. The mixture is stable for 2 weeks at -20°C and 3 days at 4°C with protection from light.
3. Hydrochloric acid as a reaction stop solution also helps to distinguish the color-difference.
4. As a standard negative control, 5 µl of G6PD-normal blood in the reaction mixture without the substrate can be used.
5. The activity can be quantified by measuring absorbance at 450-460 nm with a microplate reader. For accurate diagnosis, measurement of hemoglobin content is recommended.
6. This kit is for research use only.

1. I.S. Tantular and F. Kawamoto, Trop. Med. Int. Health, 2003, 8 (6), 569.