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Transferrin, an iron transport protein found in serum and cerebrospinal fluid, is known to be microheterogeneous with respect to its carbohydrate and sialic acid content. The forms of transferrin deficient in sialic acid and/or carbohydrate, termed carbohydrate-deficient transferrin (CDT), have been of clinical interest for almost two decades as a result of the initial finding that elevated CDT concentrations are associated with chronic, excessive alcohol abuse. We demonstrate the utility of capillary electrophoresis for examining the CDT sialoform profile via the direct electrophoresis of serum. The need for negligible preelectrophoresis sample preparation and absence of postelectrophoresis processing dramatically decreases analysis time compared to slab gel-based separations. Using a fluorocarbon-coated capillary containing a hydroxyethyl cellulose/borate buffer, the high resolution separation of serum components is effected in less than 30 min. Under these conditions, the beta region proteins (including transferrin) are well resolved from the alpha-2 and gamma zone proteins in a window where the individual transferrin sialoforms can be detected. The usefulness of this method is demonstrated with the electrophoresis of serum from subjects known to be either non-alcoholic and alcoholic.

Capillary electrophoresis represents a relatively new analytical technique. This methodology has diversified and given rise to various modes such as capillary zone electrophoresis, capillary gel electrophoresis, micellar electrokinetic capillary chromatography, capillary isoelectric focusing and capillary isotachophoresis. If capillary electrophoresis was first introduced in research laboratories, this technique is now making an entrance to the clinical laboratory. This is due to its rapid and high-efficiency separation power, its potential applications and its possible automation. Thus, capillary electrophoresis represents an attractive alternative to some time-consuming techniques. Thanks to its versatility, the use of capillary electrophoresis has been proposed for the separation and quantification of a wide spectrum of biological components ranging from macromolecules (proteins, lipoproteins, nucleic acids) to small analytes (amino acids, organic acids or drugs). This paper illustrates the potential of capillary electrophoresis which should rapidly become a major technology for a modern clinical laboratory.

We present two atypical cases of protein studies that were evaluated by immunofixation and immunosubtraction using capillary zone electrophoresis and high-resolution agarose gel electrophoresis. The first study showed an abnormal peak in the beta region by capillary zone electrophoresis that was located in the gamma region of the high-resolution agarose gel electrophoresis. Further investigation showed that this monoclonal protein was displaced due to binding with beta-lipoproteins. In the second case, a large peak was detected in the alpha-2 region and was shown by capillary zone electrophoresis to be a non-proteinaceous material that mimicked a paraprotein.

Serum protein electrophoresis and typing of monoclonal components (MCs) are routine but time-consuming and technically demanding assays. We evaluated capillary electrophoresis (Paragon CZE 2000) for automation of the two assays. CZE and cellulose acetate electrophoresis gave similar data on 794 samples. Within-run and between-run CVs were < 2% for albumin and gamma-globulins and 4-7% for alpha 1-, alpha 2-, and beta-globulins. Bilirubin, hemoglobin, triglycerides, and fibrinogen were found not to interfere. No carryover by capillaries was detected. The detection limit for MC was < 0.5 g/L. MC assessment by immunosubtraction on 403 samples identified the monoclonal type in all samples with peak concentrations > 10 g/L; only 50% of MCs that could not be quantified by densitometric scan were typed.

Capillary zone electrophoresis (CZE) offers the potential for automating serum protein electrophoretic analysis traditionally performed on standard thin-layer agarose gels. The following describes the use of CZE compared to agarose gel electrophoresis (AGE) for the detection of dysproteinemia and paraproteinemia in a clinical study involving 240 patients. The study includes within-run and between-run reproducibility data on the Paragon CZE 2000 Clinical Capillary Electrophoresis System, in addition to concordance data between the two methodologies. Paraprotein quantitation studies comparing AGE versus CZE were also performed. Reproducibility for the automated CZE system was superior to the AGE system. Improved reproducibility for the CZE method is largely due to measuring protein absorbance directly at 214 nm versus the traditional AGE method that measures the amount of dye adsorbed to protein. Reproducibility data as percent coefficient of variance (% CV) for the five classic bands in a normal control serum for between-run precision ranged from 1.2 to 4.5% for CZE compared to AGE, which ranged from 3.8 to 8.0% CV. Concordance studies between AGE and CZE involving dysproteinemias including hypogammaglobulinemia, polyclonal and monoclonal gammopathies, acute and chronic inflammation, nephrosis, hepatodegenerative disease, cirrhosis, and iron deficiency anemia showed 96% agreement. Paraprotein classification, which compared the CZE immunosubtraction method to immunofixation electrophoresis (IFE) on agarose, showed 100% agreement. Certain dysproteinemias involving beta lipoprotein were in partial concordance due to the inability of the CZE procedure to detect this component. Detection limits for monoclonal gammopathies, providing they were not comigrating with other proteins, were IgG 50 mg/dL, IgM 75 mg/dL, and IgA 75 mg/dL. Paraprotein quantitative studies between the two methods showed less than a +/- 0.2 g/dL variation.