Properties of erythrocyte light refraction in diabetic patients

Since hyperglycaemia changes the erythrocyte cell membrane fluidity and impairs cell deformity, our goal was to characterize hemoglobin and red blood cell (RBC) light refractive property changes in diabetic patients. Microscopic investigation was carried out on intact and fixed RBCs. To determine the refractive index (RI): smears of peripheral blood were air dried and fixed for 3 min in methanol. Mixtures of polyvinylpyrolidine and buffer of different pH (1:1) were used as embedding media. Intact RBCs were mixed with a buffered embedding medium, placed on a slide and overlaid with a coverslip. Interference microscopy was used for RI measurements at 18 different pH (pH = 2-13). The results showed that curves of the RI of diabetic patients and of a control group were of similar configuration, with one branch in the acidic portion of the pH scale, a maximum and two minima in the neutral (middle) portion, and one branch in the alkaline portion. The curves of the individuals from the control group overlapped each other. To the contrary, the curves of the diabetic patients were not uniform in the neutral portion and the alkaline portion. The curves of the diabetic patients in the neutral zone were shifted towards the alkaline end of the pH scale, and the RBC RI curves were lower in comparison to the control curves. The center maximum of the curves of diabetic patients corresponded to pH = 6.6 whereas the central maximum of the control group curves was at pH=6.2-6.8. Contrary to in the diabetic group, intact RBC RI curves in the control group revealed only one significantly different minimum at pH of 7.2 in the neutral zone. Using this method it is possible to show phenotypic differences between uniform type intact and fixed cells, erythrocytes of diabetic patients and of healthy donors.