Capillary zone electrophoresis for determination of carbohydrate‐deficient transferrin in human serum

Abstract

Carbohydrate-deficient transferrin (CDT) is the most specific marker for diagnosis of chronic excessive alcohol consumption and includes the serum transferrin (Tf) isoforms with two or less sialic acid residues (di-, mono-, and asialo-Tf). To monitor serum CDT, we developed a capillary zone electrophoresis (CZE) method based on the dynamic capillary coating with diethylenetriamine (DETA). The separation was performed in a bare fused-silica capillary (50 microm ID, 57 cm in length), applying a voltage of 25 kV and a temperature of 40 degrees C. Using a 100 mmol/L borate buffer, pH 8.4 with 3 mmol/L DETA, the Tf isoforms (asialo- to pentasialo-Tf) were resolved within 16 min. Enzymatic cleavage of sialic acid residues with neuraminidase and immunosubtraction were used to identify CDT isoforms. The relative amount of CDT expressed as area % of disialo-Tf isoform related to the area of tetrasialo-Tf in 50 healthy donors (24 males and 26 females; aged 25-50 years) was 3.15 +/- 0.76% (mean +/- SD). The comparison between CDT values obtained by this CZE procedure and the "Axis-Shield %CDT" kit gave r = 0.644, p < 0.001 (n = 290). This easy to use and inexpensive CZE procedure could be an ideal tool to investigate CDT proteins for clinical or forensic purposes.

Carbohydrate-deficient transferrin (CDT) as a marker of alcohol abuse: A critical review of the literature 2001–2005

“…A further dynamic coating was recently proposed by Fermo et al [30]. The reported method was similar to the previous CZE methods (separation in 100 mM borate buffer pH 8.4), but for the use of 3 mM diethylenetriamine as buffer additive.…”

Capillary electrophoresis of proteins 2003–2005

“…Besides aliphatic polyamines [41][42][43], alkylaminosilyl monomers [44] and oligoamines such as triethylenetetramine and diethylenetriamine [45,46] have also been used to reduce the surface charge and so EOF.…”

“…Carbohydrate-deficient transferrin, which is the most specific marker for the diagnosis of chronic alcoholism, was analyzed in serum by CZE [46,204]. Asialo-to pentasialo-transferrin isoforms were separated in 16 min.…”

Recent advances in the applications of CE to forensic sciences (2001–2004)

“…Most capillary electrophoretic methods are based on a zone electrophoretic separation in dynamically coated capillaries (e.g., with diaminobutane, spermine, diethylenetriamine, etc.) with direct UV detection at 200-214 nm wavelengths of the separated zones [128][129][130][131]. Quite recently, Wuyts et al [132] reported a CZE method using proprietary reagents (CEofix ® CDT buffer system, Analis, Namur, Belgium) that provide a dynamic double coating of the capillary based on polycation and polyanion.…”