Influence And Correction Of Hemolysis On Detemination Of Glucose With Glucose Oxidase Method

Objective To avoid the injury for retaking blood to patients when hemolysis occurs, plasma free hemoglobin concentration was chosed to measure the degree of hemolysis standard, double wavelengths method was adopted to determine the concentration of glucose, and to discuss the influence of hemolysis on the determination of blood sugar and the method of correction. Methods During hematolysis, the hemoglobin most affected the glucose determination, and plasma free hemoglobin concentration was chosed to measure the degree of hemolysis standard. Glucose concentrations and plasma free hemoglobin concentrations showed a certain relationship. Healthy human blood specimens were collected, without jaundice, hemolysis and lipemia. According to the document from the American Clinical and Laboratory Standards Institute (CLSI), hemoglobin solution was prepared. Hemolysis samples were made up by hemoglobin solution, plasma and calibrated solution of different concentrations of glucose in a certain proportion. Scanning200run to650nm wavelength range in the UV-VIS8500UV/visible spectrophotometer, two A-λ, curves were got, and then were compared to get the main wavelength and vice wavelength by superposition. The different hemolysis degree, determined value of glucose and real content of glucose in hemolysis samples were regarded as X, Y and Z accordingly. The experimental data was processed by SPSS13.0, and a regression equation of binary could be got. The statistical analysis of the data was processed. Finally, the relative deviation was calculated before and after regressive correction to observe whether the deviation was within permitted range. According to the United States Congress CLIA’88proficiency testing for the quality requirements of glucose analysis, the relative deviation after regressive correction within an allowable range (±10%) was judged as acceptable. And comparing regression correction and relative deviation, it was investigated whether the deviation was within an allowable range. Paired t test was observed the effect of correction. Results The determined value of glucose(Y) was lower than the real content of glucose (Z). With the deeper of the different hemolysis degree, the more obvious of the interference. The regression equation:Z=0.416+0.054X+1.015Y. Correlation coefficient r was0.974(P<0.01). Relative deviations before and after regressive correction were13.4%and7.7%respectively (P<0.01). Relative deviation after regressive correction was within permitted range (10%). Conclusions Influence of hemolysis on determination of glucose oxidase method is complex, and there are three main aspects:①Hemoglobin:the absorption spectra of hemoglobin released after red blood cells is partial overlap with quinones imine by oxidase method, showing a positive interference. Aslo, the iron heme of hemoglobin possess peroxidase activity and could catalyze a reaction of phenol and4-AAP, generatting red quinones imine, showing a positive interference.②Glutathione:Glutathione released by ruptured red blood cells can compete with hydrogen peroxide, showing a negative interference.③Dilution:Glucose concentration within the red blood cell is about20%lower than outside. The plasma (serum) is equivalent to be diluted after the red blood cells are bursted, showing a negative interference. The hemolysis has a negative influence on the method of GOD-POD, which can be corrected through the regression equation.