Separation of hemoglobin variants in single human erythrocytes by capillary electrophoresis with laser-induced native fluorescence detection.

Single human red blood cells, in which the hemoglobin (Hb) molecules exist in their native, tetrameric states, were analyzed. Upon injection and lysis of a cell, the tetramers were dissociated on-column into their respective polypeptide chains, separated, and detected by laser-induced native fluorescence detection with 275-nm excitation. This technique was applied to the determination of hemoglobin variants as found in adult (normal and elevated Hb A1) and fetal erythrocytes. Normal adult cells contained 9.6% and 4.8% glycated beta- and alpha-chains, respectively. Cells with elevated Hb A1 gave 30% and 12%, respectively. The amounts of glycated Hb and total Hb in a given cell were found to be uncorrelated.

[1]  J. L. Stafford.  Man's Hæmoglobins , 1967 .

[2]  E. Yeung,et al.  Determination of intracellular species at the level of a single erythrocyte via capillary electrophoresis with direct and indirect fluorescence detection. , 1992, Analytical chemistry.

[3]  E. Abraham,et al.  Changes in glycosylated hemoglobins with red cell aging in normal and diabetic subjects and in newborn infants of normal and diabetic mothers. , 1983, The Journal of laboratory and clinical medicine.

[4]  T. Wehr,et al.  Capillary electrophoresis of abnormal hemoglobins associated with α-thalassemias , 1993 .

[5]  J. Stuart.  The Function of Red Blood Cells: Erythrocyte Pathobiology , 1982, Progress in clinical and biological research.

[6]  M. Zhu,et al.  Capillary electrophoresis of hemoglobins and globin chains. , 1992, Journal of chromatography.

[7]  S. Fanali,et al.  Capillary zone electrophoresis and mass spectrometry for the characterization of genetic variants of human hemoglobin. , 1991, Analytical biochemistry.

[8]  R. Craver,et al.  Quantification of hemoglobin variants by capillary isoelectric focusing. , 1994, Clinical chemistry.

[9]  R. Jones,et al.  Red cell age-related changes of hemoglobins AIa+b and AIc in normal and diabetic subjects. , 1976, The Journal of clinical investigation.

[10]  D. F. Roberts,et al.  Human haemoglobin variants and their characteristics , 1977 .

[11]  P. Gallop,et al.  The glycosylation of hemoglobin: relevance to diabetes mellitus. , 1978, Science.

[12]  J. Murphy.  Influence of temperature and method of centrifugation on the separation of erythrocytes. , 1973, The Journal of laboratory and clinical medicine.

[13]  T. Satow,et al.  Effects of the sample matrix on the separation of peptides by high performance capillary electrophoresis , 1991 .

[14]  E. Yeung,et al.  Quantitative determination of native proteins in individual human erythrocytes by capillary zone electrophoresis with laser-induced fluorescence detection. , 1992, Analytical chemistry.

[15]  Douglas MacN. Surgenor,et al.  The red blood cell , 1974 .

[16]  W. Thormann,et al.  Application of dynamic capillary isoelectric focusing to the analysis of human hemoglobin variants , 1994, Electrophoresis.