Affimers as an alternative to antibodies for protein biomarker enrichment.
Protein Expr Purif
; 174: 105677, 2020 10.
Article
em En
| MEDLINE
| ID: mdl-32461183
ABSTRACT
INTRODUCTION:
Assessing the specificity of protein binders is an essential first step in protein biomarker assay development. Affimers are novel protein binders and can potentially replace antibodies in multiple protein capture-based assays. Affimers are selected for their high specificity against the target protein and have benefits over antibodies like batch-to-batch reproducibility and are stable across a wide range of chemical conditions. Here we mimicked a typical initial screening of affimers and commercially available monoclonal antibodies against two non-related proteins, IL-37b and proinsulin, to assess the potential of affimers as alternative to antibodies.METHODS:
Binding specificity of anti-IL-37b and anti-proinsulin affimers and antibodies was investigated via magnetic bead-based capture of their recombinant protein targets in human plasma. Captured proteins were analyzed using SDS-PAGE, Coomassie blue staining, Western blotting and LC-MS/MS-based proteomics.RESULTS:
All affimers and antibodies were able to bind their target protein in human plasma. Gel and LC-MS/MS analysis showed that both affimer and antibody-based captures resulted in co-purified background proteins. However, affimer-based captures showed the highest relative enrichment of IL-37b and proinsulin.CONCLUSIONS:
For both proteins tested, affimers show higher specificity in purifying their target proteins from human plasma compared to monoclonal antibodies. These results indicate that affimers are promising antibody-replacement tools for protein biomarker assay development.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proinsulina
/
Interleucina-1
/
Materiais Biomiméticos
Limite:
Humans
Idioma:
En
Revista:
Protein Expr Purif
Ano de publicação:
2020
Tipo de documento:
Article