Comparison of immunoglobulin binding capacities and ligand leakage using eight different protein A affinity chromatography matrices.
The capacities of eight protein A affinity chromatography matrices were determined in a column using a purified monoclonal mouse IgG3 antibody. The capacities ranged from 0.5 to 20 mg IgG3/ml gel. Protein A-Sepharose Fast Flow exhibited the highest capacity. Each column was tested for leakage of protein A in the presence or absence of IgG3 at pH 8.9, 4.0 and 3.0. In the absence of immunoglobulins significant leakage could only be detected at pH 8.9. In the presence of immunoglobulins, considerable leakage was also found at pH 4.0, the point at which the immunoglobulins eluted. Depending on the matrix used, the IgG3 eluted from the protein A columns exhibited a contamination of 1.8-88 ppm (weight/weight) protein A. Amongst the gels with IgG3 capacities greater than 10 mg/ml, the least contamination with protein A was observed in the IgG3 fractions from immobilized rProtein A and Protein A-Sepharose CL-4B (Fermentech).
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Sandoz AG, Basel, Switzerland.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000915:Antibody Affinity; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D005069:Evaluation Studies as Topic; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000276:immunology; D007136:Immunoglobulins / Q000276:immunology / Q000302:isolation & purification*; D008024:Ligands; D013205:Staphylococcal Protein A / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007136:Immunoglobulins; D008024:Ligands; D013205:Staphylococcal Protein A