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Glucose oxidase (GOD) converts glucose to gluconic acid. Hydrogen peroxide formed in this reaction, in presence of peroxidase (POD), oxidatively couples with 4-aminoantipyrine and phenol to produce red quinonimine dye. This dye has absorbance maximum at 505 nm. (500 - 550 nm.). The intensity of the colour complex is directly proportional to the concentration of glucose in specimen.
PREPARATION OF WORKING SOLUTION
Reconstitute enzyme & diluent as per instruction indicated on individual bottle label to prepare working solution. Mix by gentle swirling or inversion. DO NOT SHAKE VIGOROUSLY.
Phosphate Buffer : pH 7.0
For plasma separation following anticoagulants may be used :
Sodium Fluoride is preferred as anticoagulant due to its antiglycolytic activity. Higher concentration of sodium fluoride i.e. more than 10mg/ ml blood should be avoided as it may inhibit the colour development. Glucose is stable for 24 hours in neatly separated plasma and serum. If the estimation is not possible within 24 hours then the specimen should be preserved at -10 degree C and should be used within 30 days.
Serum / Plasma
Incubate the assay mixture for 15 minutes at 37 degree C or 30 minutes at room temperature (25-30 degree C). After completion of incubation period measure the absorbance against blank at 505 nm. Final colour is stable for two hours if not exposed to direct light.