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Determination of immunoglobulins in blood serum by high-performance affinity chromatography
High-performance affinity chromatography columns were prepared by immobilizing protein A from Staphylococcus aureus on 10-microns diol-bonded silica. Immunoglobulin-containing samples were injected into the column at pH 7 and eluted by stepwise changes to pH 3. Immunoglobulin G, A, and M standards were tested, but only a small fraction of the latter two was retained on the column. Analysis of 3-microliters samples of reference blood sera without pretreatment was performed using immunoglobulin G standards. Good agreement was observed between the immunoglobulin G concentrations measured by this method and by radial immunodiffusion. Chromatography of albumin and electrophoresis of retained serum fractions showed no interference by non-specific adsorption. Chromatographic analysis times were 4 min or less.
High-performance affinity chromatography columns were prepared by immobilizing protein A from Staphylococcus aureus on 10-microns diol-bonded silica. Immunoglobulin-containing samples were injected into the column at pH 7 and eluted by stepwise changes to pH 3. Immunoglobulin G, A, and M standards were tested, but only a small fraction of the latter two was retained on the column. Analysis of 3-microliters samples of reference blood sera without pretreatment was performed using immunoglobulin G standards. Good agreement was observed between the immunoglobulin G concentrations measured by this method and by radial immunodiffusion. Chromatography of albumin and electrophoresis of retained serum fractions showed no interference by non-specific adsorption. Chromatographic analysis times were 4 min or less.