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Clinical Chemistry 34: 2039-2043, 1988;
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Clinical Chemistry, Vol 34, 2039-2043, Copyright © 1988 by American Association for Clinical Chemistry

High-performance liquid affinity chromatography: rapid immunoanalysis of transferrin in serum

S Ohlson, BM Gudmundsson, P Wikstrom and PO Larsson
Perstorp Biolytica AB, Lund, Sweden.

We describe a new method for quantitatively measuring substances of clinical interest by high-performance liquid affinity chromatography (HPLAC). As a model system we selected analysis for transferrin in human serum with immobilized antibodies in a high-performance liquid chromatographic system. SelectiSpher-10 Activated Tresyl columns (5 or 10 x 0.5 cm) were used for in situ coupling of polyclonal antibodies to transferrin. The amount of transferrin eluted was determined by integrating the eluted peak at 280 nm. The whole analytical procedure-- including injection of sample, washing, elution, and analysis of data-- takes only 7 min. We characterized the HPLAC system for analysis of transferrin in several ways: intra-assay CV approximately 3%; inter- assay CV 2-9%; linear response up to 1 mg/mL column volume; detection limit approximately 3 micrograms; analytical recovery 98% +/- 2%; purity of eluted sample greater than 95% (SDS-PAGE). The HPLAC method was compared with "rocket" immunoelectrophoresis, a commonly used method of analysis for transferrin, and there was excellent correlation between the two methods (r = 0.96, n = 60). Benefits of this HPLAC technique include high precision, rapid analysis, and simplified sample handling.


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Home page
 Clin. Chem.Home page
D. S. Hage
Affinity Chromatography: A Review of Clinical Applications
Clin. Chem., May 1, 1999; 45(5): 593 - 615.
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