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Determination of Carbohydrate-deficient Transferrin Using Capillary Zone Electrophoresis

¹ Department of Clinical Chemistry, Ghent University Hospital, De Pintelaan 185, B9000 Ghent, Belgium.

² Departments of Medicine and Chemical Pathology, Medical School, The University of Zimbabwe, Avondale, Harare, Zimbabwe.

³ Department of Clinical Chemistry, Middelheim Hospital, Lindendreef 1, B2020 Antwerp, Belgium.

⁴ Department of Clinical Chemistry, AZ Stuivenberg, Lange Beeldekensstraat 267, B2060 Antwerp, Belgium.

⁵ Analis SA, 14 Rue Dewez, B5000 Namur, Belgium.
^a Address correspondence to this author at: Department of Clinical Chemistry, De Pintelaan 185, B-9000 Gent, Belgium. Fax 32-9-240-4985; e-mail joris.delanghe{at}rug.ac.be.

Background: Current methods for carbohydrate-deficient transferrin (CDT) often suffer from low precision, complexity, or risk of false positives attributable to genetic variants. In this study, a new capillary zone electrophoresis (CZE) method for CDT was developed.

Methods: CZE was performed on a P/ACE 5000 using fused-silica capillaries [50 µm (i.d.) x 47 cm] and the CEOFIX CDT buffer system with addition of 50 µL of anti-C3c and 10 µL of anti-hemoglobin. Native sera were loaded by high-pressure injection for 3 s, separated at 28 kV over 12 min, and monitored at 214 nm.

Results: CDT was completely resolved by differences in migration times (di-trisialotransferrin, 9.86 ± 0.05 min; monosialotransferrin, 9.72 ± 0.05 min; asialotransferrin, 9.52 ± 0.04 min), with a CV of 0.15%. The number of theoretical plates was 312 000 ± 21 000 for the mono- and 199 000 ± 6500 for the di-trisialylated transferrin. Genetic CB and CD variants showed prominent peaks with migration times of 10.12 ± 0.06 and 9.89 ± 0.03 min, respectively, and the carbohydrate-deficient glycoprotein syndrome could be detected, excluding false-positive results. CZE results (as a percentage; y) correlated with the Axis %CDT TIA^TM (x) values by Deming regression analysis: y = 1.92x - 7.29; r = 0.89. CDT values in 130 healthy nonalcoholics were determined. The 2.5th and 97.5th percentiles were 1.84% and 6.79%.

Conclusions: CZE without sample pretreatment can determine CDT with good precision, allows detection of variants, and correlates with ion-exchange chromatography.

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