title: AbSep--an amino acid based pseudobioaffinity adsorbent for the purification of immunoglobulin G.
abstract: The present work deals with the development and characterization of a tryptophan based pseudobioaffinity adsorbent for the purification of monoclonal and polyclonal antibodies. Tryptophan as a ligand was selected based on molecular docking and experimental screening studies of the amino acids involved in IgG-Protein A interaction. The ligand was coupled to a polymethacrylate based rigid, porous SEPABEADS beaded matrix to obtain the desired affinity adsorbent, which was named AbSep. Characterization studies with regards to the effect of matrix properties (pore size, particle size, nature of matrix, spacer arm) and the medium properties (pH, conductivity, additives) were performed to elucidate the nature of IgG-AbSep interactions and to determine the optimal conditions for obtaining high binding and purity of IgG. The equilibrium binding capacity of AbSep and dissociation constant was found to be 78 mg/ml and 5.31×10(-6)M respectively. AbSep was able to successfully purify polyclonal human IgG from plasma and monoclonal antibody (chimeric IgG1) from CHO cell culture supernatant. Both binding and elution steps were performed at near neutral pH resulting in a purity and recovery of more than 90% and 85% respectively. Additionally, AbSep was shown to be stable to 0.5M NaOH solutions, the preferred agent for cleaning and sanitization of chromatographic media.
authors.affiliation: Department of Chemical Engineering, Institute of Chemical Technology, University of Mumbai, N. M. Parekh Marg, Matunga, Mumbai, India. adnaik2@ncsu.edu
authors.forename: Amith D
authors.initials: AD
authors.lastname: Naik
authors.forename: Monika
authors.initials: M
authors.lastname: Raina
authors.forename: Arvind M
authors.initials: AM
authors.lastname: Lali
mesh_terms: D024322:Amino Acids, Aromatic / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification* / Q000378:metabolism; D016466:CHO Cells; D002846:Chromatography, Affinity / Q000379:methods*; D006224:Cricetinae; D003412:Cricetulus; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D010316:Particle Size; D011092:Polyethylene Glycols; D016062:Porosity; D011485:Protein Binding; D012965:Sodium Chloride; D013205:Staphylococcal Protein A / Q000737:chemistry / Q000378:metabolism
chemical_list: D024322:Amino Acids, Aromatic; D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D013205:Staphylococcal Protein A; D011092:Polyethylene Glycols; D012965:Sodium Chloride