title: Development of a high throughput protein a well-plate purification method for monoclonal antibodies.
abstract: We have developed a new high throughput method for the purification of monoclonal antibodies from harvested cell culture fluid for analytical characterization. This method uses Protein A resin in a 96 well-plate format with protein loading sufficient to perform multiple analyses per well. Resin and buffer conditions were optimized to obtain aggregate and charge variant comparability with three preparative Protein A purified monoclonal antibodies. We are able to successfully demonstrate comparability for aggregate within 0.25% based upon size-exclusion chromatography. Acidic species were found to be within 2% from the preparative purified control based upon cation-exchange chromatography, 5% based upon capillary zone electrophoresis, and 3% based upon imaged capillary isoelectric focusing. Glycan distribution was analyzed and was within 1% of the preparative purified controls. A tryptic digest was performed and all peaks in the preparative purified control were found in the first elution from the well-plate format.
authors.affiliation: Genentech Inc., Oceanside, CA 92056, USA. hopp.jennifer@gene.com
authors.forename: Jennifer
authors.initials: J
authors.lastname: Hopp
authors.forename: Ross
authors.initials: R
authors.lastname: Pritchett
authors.forename: Maria
authors.initials: M
authors.lastname: Darlucio
authors.forename: Junfen
authors.initials: J
authors.lastname: Ma
authors.forename: Judy H
authors.initials: JH
authors.lastname: Chou
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000295:instrumentation / Q000379:methods*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000302:isolation & purification; D007475:Ion Exchange Resins; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D013205:Staphylococcal Protein A / Q000737:chemistry
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007475:Ion Exchange Resins; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A