Study of the feasibility of using a pellicular anion-exchange column for separation of transferrin isoforms in human serum by HPLC with UV detection

A method has been optimised for the separation of glycoforms of human serum transferrin, using a high-performance pellicular anion-exchange chromatographic column. The effect of the eluent pH and of the column temperature on the separation of transferrin glycoforms was studied using a standard solution of commercially available human serum transferrin. An HPLC system equipped with an ultraviolet detector was used for the analysis. No immunoassay was used after the anion-exchange chromatographic separation of the glycoforms, in contrast with most currently used methods. The method was applied to the separation and quantification of transferrin glycoforms in serum from a healthy, non-pregnant woman, after saturation of transferrin with iron and further precipitation of lipoproteins. The whole chromatographic run, including re-equilibration of the column, took 35 min.