Comparative study of nitro blue tetrazolium (NBT) reduction method for estimation of glycated haemoglobin with glycated HbA1c estimated on DCA2000+analyzer (immunoagglutination inhibition).

Glycated haemoglobin is a diagnostic tool, used for the monitoring of the glycemic status among diabetic patients. The present study is designed to compare and correlate modified NBT reduction method for the estimation of Glycated protein (Glycated Haemoglobin) with HbA1C estimated on DCA+2000Analyzer. Glycated protein reduces Nitro Blue Tetrazolium (NBT) reagent in alkaline medium to tetrazinolyl radical NBT+ which is disproportional to yield a highly colored formazan dye (MF+) (monoformazen), absorbance of colored compound was measured which gives the concentration of glycated proteins present in the sample. Heme free globin (glycated hemoglobin) was extracted out and dissolved immediately in 1 ml normal saline. Dissolved globin was treated with modified NBT reagent, absorbance of color developed was recorded in milli delta A/min. The results of modified NBT were then compared with HbA1c estimated by immunoagglutination inhibition method. Correlation coefficient between Glycated hemoglobin and HbAlc was found to be r=0.926 using Schimadzu CL-750 spectrophotometer and r=0.902 using colorimeter. Results of this study were found to be statistically significant p < 0.001. Thus the present study concludes that Glycated hemoglobin testing by modified NBT reduction method is as sensitive as HbAlc estimated by DCA2000+Analyzer (immunoagglutination inhibition). Hence it could be used for routine monitoring of blood glucose control level in diabetic subjects.