Monitoring of transferrin isoforms in biological samples by capillary electrophoresis

Abstract

Work dealing with the monitoring of transferrin isoforms in human serum and other body fluids by capillary electrophoresis is reviewed. It comprises capillary zone electrophoresis and capillary isoelectric focusing efforts that led to the exploration and use of assays for the determination of carbohydrate‐deficient transferrin as a marker for excessive alcohol intake, genetic variants of transferrin, congenital disorders of glycosylation and β‐2‐transferrin, which is a marker for cerebrospinal fluid leakage. This paper provides insight into the development, specifications, strengths, weaknesses, and routine use of the currently known capillary electrophoresis based assays suitable to detect transferrin isoforms in body fluids. The achievements reached so far indicate that capillary zone electrophoresis is an attractive technology to monitor the molecular forms of transferrin in biological specimens as the assays do not require an elaborate sample pretreatment and thus can be fully automated for high‐throughput analyses on multicapillary instruments. Assays based on capillary isoelectric focusing are less attractive. They require immunoextraction of transferrin from the biological matrix and mobilization after focusing if instrumentation with a whole‐column imaging detector is not available. Interactions of the carrier ampholytes with the iron of transferrin may prevent iron saturation and thus provide more complicated isoform patterns.

High‐resolution capillary zone electrophoresis for transferrin glycoform analysis associated with congenital disorders of glycosylation

“…The electropherogram of Figure B does not reveal the presence of an asialo‐Tf that could be quantified. This is somewhat different compared to the sera of alcohol abusers for which an asialo‐Tf peak is typically monitored when the disialo‐Tf peak is larger than 3.0% . Migration times of the Tf isoforms monitored in panels 3A and B were identical.…”

“…This Tf profile has a higher level of disialo‐Tf compared to trisialo‐Tf, which is different than in the control sample used (Figure A). The disialo‐Tf levels in the sera of alcohol abusers can be lower, equal, or higher than trisialo‐Tf . Thus, the ALG13‐CDG pattern shown in Figure A cannot be distinguished from that of an alcohol abuser.…”

“…It is interesting to note that large and broad trisialo‐Tf peaks that can even overlap the disialo‐Tf peak are occasionally observed for sera of patients with liver cirrhosis. This situation is referred to as disialo‐trisialo‐Tf bridging and was reported to be a result of increased branching and fucosylation of the carbohydrate moiety . Furthermore, sera of PMM2‐CDG patients also contain small amounts of monosialo‐Tf (Figure ).…”

“…Following excessive alcohol ingestion for extended time periods, the amounts of less glycosylated forms of Tf (disialo-Tf > 2%, monosialo-Tf, and asialo-Tf) are increasing. Less glycosylated forms of Tf are also found in sera of CDG patients [23,24,[28][29][30][31][32][33][34], CSF [24], and can be produced in vitro by addition of neuraminidase to serum [16].…”

“…Tf isoform analysis by CZE is performed with automated multicapillary platforms (CAPILLARYS of Sebia, Evry, France) or with the CDT reagent kit CEofix (Analis, Namur, Belgium) on single capillary instruments of different manufacturers [23,24]. The latter system comprises polymeric polyions that form a dynamic double coating, an approach that leads to highly reproducible data [16], and was optimized for high-resolution analyses in our laboratory [17], which is advantageous for analysis of patient sera with unusual Tf patterns, including those with high amounts of trisialo-Tf and genetic variants [23][24][25][26][27]. CDG serum Tf patterns using the CEofix approach [11,23,28,29], the CAPILLARYS assay [30,31], and laboratory-made assays [32][33][34][35] have been reported.…”