Quantitative analysis of multiple molecular forms of transferrin using isoelectric focusing and zone immunoelectrophoresis assay (ZIA)

Abstract

A new technique for the quantitative determination of electrophoretically separated proteins has been developed. The technique has been used for quantification of a molecular form of human serum transferrin having an isoelectric point (pI) of 5.7. The proteins in human serum samples were separated by isoelectric focusing in agarose gel. With guidance by a coloured pI-marker protein (i.e., carbamylated horse muscle myoglobin), gel pieces containing the transferrin (pI 5.7) were isolated, disintegrated with a syringe, and deposited directly as samples in zone immunoelectrophoresis assay (ZIA). The transferrin content of the gel pieces was determined with a relative standard deviation (R.S.D.) of 7%. The total transferrin concentration in the serum samples was also determined by ZIA with an R.S.D. of 3%. The technique appears applicable to the determination of many other proteins.