Improved capillary electrophoresis determination of carbohydrate-deficient transferrin including on-line immunosubtraction

Abstract

The instrumental analysis of carbohydrate-deficient transferrin (CDT), a recognized marker of chronic alcohol abuse, is most commonly carried out by high-performance liquid chromatography (HPLC) or capillary zone electrophoresis (CZE). Between these two techniques, CZE shows higher efficiency and productivity, but is often reported to be inferior to HPLC in terms of selectivity, because of a less specific ultraviolet detection wavelength than HPLC. On these grounds, the present work was aimed at the development of an improved CZE method for CDT determination, including an on-line immunosubtraction step specifically aimed at enhancing the analytical specificity of CZE determination. The analytical conditions were as follows: uncoated fused silica capillary, 30 microm x 60 cm (L = 50 cm to detector); running buffer, 100 mmol/L borate and 6 mmol/L DAB (1,4-diaminobutane), pH 8.3; voltage, 30 kV; temperature, 25 degrees C; detection, 200 nm. Under the described CZE conditions, a baseline separation between all the CDT related peaks was achieved with good analytical performances in terms of both precision and accuracy. In order to achieve unequivocal recognition of the CDT peaks, an in-capillary immunosubtraction step was included by loading a plug of anti-human transferrin antibody solution after the sample plug. This analytical approach was applied successfully to recognize CDT peaks in the presence of potential interferences.