Effect of cationic buffer additives on the capillary electrophoretic separation of serum transferrin from different species

The presence of specific transferrin (Tf) glycoforms inhuman serum has been shown to correlate with certain clinical syndromes. Hence, the ability to separate and quantitatively measure the various forms of human transferrin has become increasingly important. As a means of evaluating the potential for developing a rapid capillary electrophoresis-based assay for the analysis of carbohydrate-deficient transferrins (CDTs), the capillary zone electrophoretic (CZE) analysis of Tfs from several species was evaluated using uncoated capillaries and a separation augmented with cationic additives. With bovine Tf, five peaks (representing different sialylated forms) were partially resolved in berate and baseline-resolved when 1,4-diaminobutane was added to the buffer. These same conditions were found to be inadequate for the resolution of the sialoforms from other species. Some success was achieved using alpha,omega-bis-quaternary ammonium alkanes instead of the 1,4-diaminobutane and optimizing the pH for each of the species Tfs. Human Tf was found to be resolved in an uncoated capillary equilibrated with a berate buffer containing millimolar concentrations of decamethonium bromide as a buffer additive. Under these conditions, resolution of the various sialoforms from the iron-saturated Tf was possible and the glycoforms were found to migrate differently than their iron-depleted counterparts. Despite the resolution achievable under these conditions, the lengthy analysis time is incompatible with the requirements for a clinical CZE-based assay.