title: Antibody purification: affinity chromatography - protein A and protein G Sepharose.
abstract: Affinity chromatography relies on the reversible interaction between a protein and a specific ligand immobilized in a chromatographic matrix. The sample is applied under conditions that favor specific binding to the ligand as the result of electrostatic and hydrophobic interactions, van der Waals' forces and/or hydrogen bonding. After washing away the unbound material the bound protein is recovered by changing the buffer conditions to those that favor desorption. The technique has been used not only to isolate antigen-specific antibodies but also to remove specific contaminants from biological samples. Methods are described for the purification of immunoglobulins, namely IgG, IgG fragments and subclasses, using the high affinity of protein A and protein G coupled to agarose. In the Subheading 3 there are also protocols for affinity purification using a specific ligand coupled to commercial matrices like CNBr- Sepharose 4-B and Affigel.
authors.affiliation: Receptors and Signal Transduction Section, Oral Infection and Immunity Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Ana Cristina
authors.initials: AC
authors.lastname: Grodzki
authors.forename: Elsa
authors.initials: E
authors.lastname: Berenstein
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods*; D006801:Humans; D007074:Immunoglobulin G / Q000302:isolation & purification; D008024:Ligands; D009419:Nerve Tissue Proteins / Q000737:chemistry; D012685:Sepharose / Q000737:chemistry; D013205:Staphylococcal Protein A / Q000737:chemistry
chemical_list: D000906:Antibodies; C029152:G-substrate; D007074:Immunoglobulin G; D008024:Ligands; D009419:Nerve Tissue Proteins; D013205:Staphylococcal Protein A; D012685:Sepharose