SEMIAUTOMATIC METHOD FOR DETERMINATION OF DIFFERENT ISOFORMS OF CARBOHYDRATE-DEFICIENT TRANSFERRIN

Carbohydrate deficient transferrin (CDT) has been reported to be one o f the best biochemical markers of alcohol abuse. However, a need still exists for a simple and practical method for widespread laboratory us e. A semi-automatic (SA) isoelectric focusing (IEF) assay for CDT (SA- IEF-CDT) by a Phast System(TM) is introduced here. Different isoforms of transferrin were separated by IEF on polyacrylamide gels (pI 4.0-6. 5) and located by immunofixation with an anti-transferrin serum. The p recipitation bands were stained with Coomassie Brilliant Blue and quan titated densitometrically. The present method gave a picture of the re lative amounts of 10 different transferrin isoforms. The percentage of CDT with pI greater-than-or-equal-to 5.7 (representing di-, mono- and asialotransferrin) was calculated. For comparisons transferrin bands with pI greater-than-or-equal-to 5.6 (tri-, di-, mono-, and asialotran sferrin), pI greater-than-or-equal-to 5.8 (mono- and asialotransferrin ) and pI greater-than-or-equal-to 5.9 (asialotransferrin) as well as G GT, ASAT and ALAT were calculated. The method showed good linearity an d it identified different isoforms in concentrations of < 10 mg/l of t ransferrin. The correlation of the present method with a commercially available method employing anion exchange followed by double antibody RIA (AE-RIA-CDT) was good (n = 38, r = 0.924). In 19/20 (95%) of healt hy controls, the CDT value was below 4.45 (mean + 2 S.D.) of total tra nsferrin, while higher values were observed in all 20 (100%) alcoholic s. In conclusion, the developed semi-automatic method is a practical a nd reliable alternative for determination of different transferrin iso forms.