Purification of antibody fragments via interaction with detergent micellar aggregates.
Sci Rep
; 11(1): 11697, 2021 06 03.
Article
em En
| MEDLINE
| ID: mdl-34083598
ABSTRACT
The research described in this report seeks to present proof-of-concept for a novel and robust platform for purification of antibody fragments and to define and optimize the controlling parameters. Purification of antigen-binding F(ab')2 fragments is achieved in the absence of chromatographic media or specific ligands, rather by using clusters of non-ionic detergent (e.g. Tween-60, Brij-O20) micelles chelated via Fe2+ ions and the hydrophobic chelator, bathophenanthroline (batho). These aggregates, quantitatively capture the F(ab')2 fragment in the absence or presence of E. coli lysate and allow extraction of only the F(ab')2 domain at pH 3.8 without concomitant aggregate dissolution or coextraction of bacterial impurities. Process yields range from 70 to 87% by densitometry. Recovered F(ab')2 fragments are monomeric (by dynamic light scattering), preserve their secondary structure (by circular dichroism) and are as pure as those obtained via Protein A chromatography (from a mixture of F(ab')2 and Fc fragments). The effect of process parameters on Ab binding and Ab extraction (e.g. temperature, pH, ionic strength, incubation time, composition of extraction buffer) are reported, using a monoclonal antibody (mAb) and polyclonal human IgG's as test samples.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteína Estafilocócica A
/
Fragmentos Fab das Imunoglobulinas
/
Fragmentos de Imunoglobulinas
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article