Concanavalin A crossed affinity immunoelectrophoresis and image analysis for semiquantitative evaluation of microheterogeneity profiles of human serum transferrin from alcoholics and normal individuals

Heegaard, Niels H H; Hagerup, Martin; Thomsen, Åge Chr.; Heegaard, Peter M. H.

doi:10.1002/elps.1150101207

Cited by 22 publications

(6 citation statements)

References 29 publications

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“…All require presaturation with iron to reduce microheterogeneity. Initially, lectin affinity crosselectrophoresis [99] and IEF followed by immunoblotting [95] were used to separate, identify, and quantify the isoforms. Currently, chromatographic tests based on anion exchange, or IEF are used to fractionate the CDT isoforms.…”

Section: Carbohydrate-deficient Transferrins (Tf's)mentioning

confidence: 99%

Capillary electrophoresis and the clinical laboratory

et al. 2006

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Over the past 15 years, CE as an analytical tool has shown great promise in replacing many conventional clinical laboratory methods, such as electrophoresis and HPLC. CE's appeal was that it was fast, used very small amounts of sample and reagents, was extremely versatile, and was able to separate large and small analytes, whether neutral or charged. Because of this versatility, numerous methods have been developed for analytes that are of clinical interest. Other than molecular diagnostic and forensic laboratories CE has not been able to make a major impact in the United States. In contrast, in Europe and Japan an increasing number of clinical laboratories are using CE. Now that automated multicapillary instruments are commercially available along with cost-effective test kits, CE may yet be accepted as an instrument that will be routinely used in the clinical laboratories. This review will focus on areas where CE has the potential to have the greatest impact on the clinical laboratory. These include analyses of proteins found in serum and urine, hemoglobin (A1c and variants), carbohydrate-deficient transferrin, forensic and therapeutic drug screening, and molecular diagnostics.

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Section: Carbohydrate-deficient Transferrins (Tf's)mentioning

confidence: 99%

Capillary electrophoresis and the clinical laboratory

et al. 2006

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“…This fact concluded that the fucosylated Tf in highly branched glycans increased in HACTf molecules. The structural heterogeneity of the carbohydrate moiety of human serum Tf has been reported in some physicopathologic conditions [19,22,23] as well as in vitro hepatoma cell line [24]. Fucosylation of the carbohydrate moiety of the proteins has been demonstrated in some proteins, such as Tf [24], a-fetoprotein [25] and y-glutamyl transpeptidase [26].…”

Section: Discussionmentioning

confidence: 99%

Clear cell carcinoma of the human ovary synthesizes and secretes a transferrin with microheterogeneity of lectin affinity

et al. 1990

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Human ovarian clear cell carcinoma cell line (transferrin (Tf)-non-producer), HAC 2, cells were adapted to grow in chemically defined synthetic medium when the cells were cultured with medium containing 10 pg/ml of insulin at least for 6 months. They synthesized and secreted constantly the 80 kDa protein immunologically similar to human serum Tf (15 f 12 ng/ml/lO' cells/3 days). By sensitive lectin-affinity electrophoresis followed by antibody-affinity blotting technique, a concanavalin A weakly bound or unbound, lentil lectin, a strongly reactive abnormal band, which was rarely found in human serum Tf, was detectable in the Tf synthesized by HAC 2 cells (HACTf). These fmdings suggest that the HACTf may act as one of the autocrine growth factors and that this heterogeneity of HACTf for lectin affinity is ascribed to differences in the carbohydrate moiety of the Tf.

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“…Early procedures for CDT determination were based on concanavalin A crossed affinity immunoelectrophoresis [6] and on IEF coupled with laser densitometry to evaluate T f band patterns after immunoblotting [7]. The latter highly selective technique, in spite of its complexity for routine clinical use, is considered the reference method for the diagnosis of congenital disorders of glycosylation.…”

Section: Introductionmentioning

confidence: 99%

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

et al. 2004

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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.

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Concanavalin A crossed affinity immunoelectrophoresis and image analysis for semiquantitative evaluation of microheterogeneity profiles of human serum transferrin from alcoholics and normal individuals

Cited by 22 publications

References 29 publications

Capillary electrophoresis and the clinical laboratory

Capillary electrophoresis and the clinical laboratory

Clear cell carcinoma of the human ovary synthesizes and secretes a transferrin with microheterogeneity of lectin affinity

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

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