Analysis of Carbohydrate-Deficient Transferrin: Comparative Evaluation of Turbidimetric Immunoassay, Capillary Zone Electrophoresis, and HPLC

Abstract

Most assays for carbohydrate-deficient transferrin (CDT) (1)(2)(3) use cartridge extraction of CDT isoforms followed by immunoassay (4)(5). In 2001 the US Food and Drug Administration cleared the %CDT turbidimetric immunoassay (TIA; Axis Shield Plc) for detection of sustained and harmful alcohol use. Another method, based on anion-exchange HPLC separation of the CDT isoforms with direct detection at 460 nm [selective for the iron–transferrin (Tf) complex], was first developed by Jeppsson et al. (6) and adopted with minor changes by several authors, who reported advantages over immunoassays in terms of analytical selectivity, accuracy, and precision (7)(8)(9)(10). Recently released commercial reagents for HPLC analysis (Recipe®) offer advantages in interlaboratory analytical standardization. Capillary zone electrophoresis (CZE) with dynamically coated capillaries and direct ultraviolet detection at 200 nm (11)(12)(13)(14)(15)(16)(17) has also been successfully applied to CDT determination in human serum (2), and a CZE method with a proprietary capillary coating is commercially available (CEofix® CDT; Analis). Validation studies for each of these techniques have been reported in the literature, but no direct comparison studies have been published. We compared 3 commercial methods, based on TIA, CZE, and HPLC, focusing on their application to the diagnosis of chronic alcohol abuse in the forensic environment and in other areas requiring high diagnostic reliability and objectivity. CDT immunoassays were performed with a commercial reagent set (%CDT TIA) from Axis Shield on a BN2 nephelometer (Dade Behring), according to the manufacturer’s instructions. The procedure included iron saturation followed by CDT extraction onto anion-exchange disposable cartridges and a TIA. CZE was performed on a P/ACE MDQ capillary electropherograph (Beckman Coulter) with uncoated fused-silica capillaries [50 cm (total length) × 50 μm (i.d.)] obtained from Polymicro …