title: Hydroxyapatite chromatography: purification strategies for recombinant proteins.
abstract: The purpose of this section is to provide developmental purification strategies for recombinant proteins on a column of hydroxyapatite and then to optimize them for repeated purification cycles using effect additives. Surface neutralization solutions (SNS) (patent pending) and/or the addition of calcium ions reduce the solubility of hydroxyapatite thus extending the column lifetime. This section leads the user through purification protocol screening followed by SNS--calcium ion supplementation step gradients for purifying a monoclonal antibody from its contaminants. The section provides simple analytical tools using a 160 ml scale column to predict column life at the process scale. However, the development engineer is encouraged to test the model in a minimum-sized process column.
authors.affiliation: Bio-Rad Laboratories, Inc., Hercules, CA, USA. Electronic address: larry_cummings@bio-rad.com.
authors.forename: Larry J
authors.initials: LJ
authors.lastname: Cummings
mesh_terms: D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D002118:Calcium / Q000032:analysis; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D017886:Durapatite; D004867:Equipment Design; D006863:Hydrogen-Ion Concentration; D011994:Recombinant Proteins / Q000302:isolation & purification*
chemical_list: D000911:Antibodies, Monoclonal; D011994:Recombinant Proteins; D017886:Durapatite; D002118:Calcium
keywords: Calcium ion analysis; Hydroxyapatite chromatography; Phosphoryl groups; Purification screening protocol; SEC profile; Surface neutralization solutions (SNS); mAb fraction; pH measurement