Affinity purification of mouse monoclonal IgE using a protein A mimetic ligand (TG19318) immobilized on solid supports

A synthetic ligand (TG19318), deduced from the screening of a combinatorial peptide library, has been previously characterized by our group for its applicability in affinity chromatography for polyclonal and monoclonal IgG purification from crude sources. In this study we have extended the characterization of its recognition properties for other immunoglobulin classes, evaluating its ability to purify mouse monoclonal IgE from ascitic fluid. TG19318 affinity columns proved useful for a very convenient one-step purification of IgE directly from crude ascites, by loading the samples on the columns equilibrated with 50 mM sodium phosphate at pH 7 and eluting and adsorbed IgE by a buffer change to 0.1 M acetic acid. Antibody purity after affinity purification was very high and no albumin traces were detected, as determined by SDS-PAGE analysis. Antibody activity was fully recovered after purification, as determined by immunoassays on antigen-coated plates, and up to 5 mg of IgEs could be purified on a 1 ml column in a single run.