title: Ligand affinity chromatographic separation of serum IgG on recombinant protein G-silica.
abstract: We purified IgG by an automated chromatographic procedure, using a recombinant DNA-produced Protein G ligand bound to silica as the solid phase. The IgG can be separated from human serum in approximately 20 min with little operator manipulation. Recovery studies indicate that unconcentrated eluates contain 77-100% of the applied IgG. Concentration to volumes approximating that of the applied serum resulted in recovery of 76-98% of the applied IgG. High-resolution protein electrophoresis of the eluates demonstrated retention of the broad gamma band, with virtual absence of other serum proteins. This finding was confirmed by immunoelectrophoresis, which demonstrated homogeneity of IgG in the eluates. IgG subclass distributions in four eluates, compared with the sera from which they were harvested, indicate comparable percentages of IgG1, IgG2, and IgG3 and some enhancement of the IgG4 fraction [serum IgG4: 2.9 +/- 0.4% (SEM); eluate IgG4: 13.8 +/- 1.7%, P = 0.007]. The reactivity of eluates from two sera containing various antibodies to human immunodeficiency virus type 1 was virtually identical to that of the sera from which they were prepared. We conclude that this chromatographic application is an effective purification method for serum IgG and antibodies of the IgG class. The method may be applicable in the specialty clinical laboratory, particularly in those interested in protein and other immunological abnormalities.
authors.affiliation: Department of Medicine, Seton Hall University, School of Graduate Medical Education.
authors.forename: P
authors.initials: P
authors.lastname: Cassulis
authors.forename: M V
authors.initials: MV
authors.lastname: Magasic
authors.forename: V A
authors.initials: VA
authors.lastname: DeBari
mesh_terms: D002846:Chromatography, Affinity; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D007122:Immunoelectrophoresis / Q000379:methods; D007074:Immunoglobulin G / Q000302:isolation & purification*; D009101:Multiple Myeloma / Q000097:blood / Q000276:immunology; D011994:Recombinant Proteins
chemical_list: D007074:Immunoglobulin G; D011994:Recombinant Proteins