Document Detail


A dual-mode approach to the selective separation of antibodies and their fragments.
MedLine Citation:
PMID:  11393728     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
A novel chromatography method for the separation of antibodies is described. The adsorption of antibodies on the solid phase involves interaction with a ligand that combines mild hydrophobic characteristics and some degree of molecular recognition with a derivative of pyridine. This combined effect results in the adsorption of antibodies in the absence of lyotropic salts. When environmental pH is changed, the ligand becomes ionically charged, allowing the desorption of antibodies. The mechanism of adsorption, involving hydrophobic associations and ionic related interaction, is here qualified as dual-mode. Studies on the determination of the apparent dissociation constant for immunoglobulins G are presented. Adsorption of antibodies from crude feedstocks typically occurs without adjustment of pH or ionic strength. The sorbent is then washed with a buffer to eliminate protein impurities and, when lowering the environmental pH, antibodies are desorbed. The solid-phase material is used for the separation of antibodies from an ascites fluid and from a cell culture supernatant, followed by a polishing step on an hydroxyapatite column. Preliminary studies, related to the ability of the solid phase to separate antibody fragments, are also reported. In these studies, it has been demonstrated that both Fab and Fc fragments from polyclonal IgG are adsorbed to the solid phase under typical binding conditions. Under other defined physico-chemical conditions (ionic strength and pH), separation of both fragments in a single step has been achieved.
Authors:
L Guerrier; I Flayeux; E Boschetti
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Journal of chromatography. B, Biomedical sciences and applications     Volume:  755     ISSN:  1387-2273     ISO Abbreviation:  J. Chromatogr. B Biomed. Sci. Appl.     Publication Date:  2001 May 
Date Detail:
Created Date:  2001-06-06     Completed Date:  2001-10-04     Revised Date:  2005-11-17    
Medline Journal Info:
Nlm Unique ID:  9714109     Medline TA:  J Chromatogr B Biomed Sci Appl     Country:  Netherlands    
Other Details:
Languages:  eng     Pagination:  37-46     Citation Subset:  IM    
Affiliation:
Biosepra S.A., Life Technologies, Cergy-Pontoise, France. lguerrie@lifetech.com
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MeSH Terms
Descriptor/Qualifier:
Adsorption
Antibodies / isolation & purification*
Ascitic Fluid / chemistry,  immunology
Chromatography / methods
Chromatography, High Pressure Liquid
Culture Media / chemistry
Humans
Hydrogen-Ion Concentration
Hydroxyapatites
Immunoglobulin Fab Fragments / isolation & purification
Immunoglobulin Fc Fragments / isolation & purification
Immunoglobulin Fragments / isolation & purification*
Immunoglobulin G / chemistry,  isolation & purification
Ligands
Osmolar Concentration
Chemical
Reg. No./Substance:
0/Antibodies; 0/Culture Media; 0/Hydroxyapatites; 0/Immunoglobulin Fab Fragments; 0/Immunoglobulin Fc Fragments; 0/Immunoglobulin Fragments; 0/Immunoglobulin G; 0/Ligands

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