Capillary zone electrophoresis for determination of carbohydrate-deficient transferrin in human serum.
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.
Fermo
Isabella
I
Laboratory of Separative Techniques, IRCCS H San Raffaele, Milan, Italy.
Germagnoli
Luca
L
Soldarini
Armando
A
Dorigatti
Fernanda
F
Paroni
Rita
R
D000328:Adult; D000437:Alcoholism / Q000175:diagnosis*; D015415:Biomarkers / Q000097:blood; D002241:Carbohydrates / Q000172:deficiency; D019075:Electrophoresis, Capillary / Q000379:methods* / Q000592:standards; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009439:Neuraminidase / Q000378:metabolism; D011073:Polyamines; D012794:Sialic Acids / Q000378:metabolism; D014168:Transferrin / Q000032:analysis* / Q000737:chemistry
D015415:Biomarkers; D002241:Carbohydrates; D011073:Polyamines; D012794:Sialic Acids; D014168:Transferrin; C005391:diethylenetriamine; D009439:Neuraminidase