title: Minibodies and Multimodal Chromatography Methods: A Convergence of Challenge and Opportunity.
abstract: This case study describes early phase purification process development for a recombinant anticancer minibody produced in mammalian cell culture. The minibody did not bind to protein A. Cation-exchange, anion-exchange, hydrophobic-interaction, and hydroxyapatite (eluted by phosphate gradient) chromatographic methods were scouted, but the minibody coeluted with BSA to a substantial degree on each. Hydroxyapatite eluted with a sodium chloride gradient separated BSA and also removed a dimeric contaminant, but BSA consumed so much binding capacity that this proved impractical as a capture tool. Capto MMC media proved capable of supporting adequate capture and significant dimer removal, although both loading and elution selectivity varied dramatically with the amount of supernatant applied to the column. An anion-exchange step was included to fortify overall virus and DNA removal. These results illustrate the value of multimodal chromatography methods when affinity chromatography methods are lacking and conventional alternatives prove inadequate.
authors: Pete Gagnon is principal consultant at Validated Biosystems Inc., 240 Avenida Vista Montana, Suite 7F, San Clemente, CA USA 92672; 1-949-276-7477, fax 1-949-606-1904;  www.validated.com . Chia-Wei Cheung, Mark A. Sherman, Andrew A. Raubitschek, and Paul J. Yazaki are with the department of cancer immunotherapeutics and tumor immunology at the City of Hope Medical Center's Beckman Research Institute in Duarte, CA;  www.cityofhope.org/research/beckman-research-institute . Eric J. Lepin and Anna M. Wu are with the Crump Institute for Molecular Imaging's department of molecular and medical pharmacology at UCLA's David Geffen School of Medicine in Los Angeles, CA;  www.crump.ucla.edu.
authors: Pete
authors: P
authors: Gagnon
authors: Chia-Wei
authors: CW
authors: Cheung
authors: Eric J
authors: EJ
authors: Lepin
authors: Anna M
authors: AM
authors: Wu
authors: Mark A
authors: MA
authors: Sherman
authors: Andrew A
authors: AA
authors: Raubitschek
authors: Paul J
authors: PJ
authors: Yazaki