Determination of carbohydrate-deficient transferrin using capillary zone electrophoresis.
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 microm (i.d.) x 47 cm] and the CEOFIX CDT buffer system with addition of 50 microL of anti-C3c and 10 microL 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 (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.
Wuyts
B
B
Department of Clinical Chemistry, Ghent University Hospital, De Pintelaan 185, B9000 Ghent, Belgium.
Delanghe
J R
JR
Kasvosve
I
I
Wauters
A
A
Neels
H
H
Janssens
J
J
D000328:Adult; D000368:Aged; D002852:Chromatography, Ion Exchange; D019075:Electrophoresis, Capillary; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D020033:Protein Isoforms / Q000032:analysis / Q000235:genetics; D012016:Reference Values; D012680:Sensitivity and Specificity; D014168:Transferrin / Q000031:analogs & derivatives* / Q000032:analysis* / Q000235:genetics
D020033:Protein Isoforms; D014168:Transferrin; C056759:carbohydrate-deficient transferrin