On-line bioaffinity-electrospray mass spectrometry for simultaneous detection, identification, and quantification of protein-ligand interactions.
J Am Soc Mass Spectrom
; 21(10): 1643-8, 2010 Oct.
Article
em En
| MEDLINE
| ID: mdl-20692851
ABSTRACT
We describe here an on-line combination of a surface acoustic wave (SAW) biosensor with electrospray ionization mass spectrometry (SAW-ESI-MS) that enables the direct detection, identification, and quantification of affinity-bound ligands from a protein-ligand complex on a biosensor chip. A trapping column was used between the SAW-biosensor and the electrospray mass spectrometer equipped with a micro-guard column, which provides simultaneous sample concentration and desalting for the mass spectrometric analysis of the dissociated ligand. First applications of the on-line SAW-ESI-MS combination include (1), differentiation of ß-amyloid (Aß) epitope peptides bound to anti-Aß antibodies; (2), the identification of immobilized Substance P peptide-calmodulin complex; (3), identification and quantification of the interaction of 3-nitrotyrosine-modified peptides with nitrotyrosine-specific antibodies; and (4), identification of immobilized anti-α-synuclein-human α-synuclein complex. Quantitative determinations of protein-ligand complexes by SAW yielded dissociation constants (K(D)) from micro-to low nanomolar sample concentrations. The on-line bioaffinity-ESI-MS combination presented here is expected to enable broad bioanalytical application to the simultaneous, label-free determination and quantification of biopolymer-ligand interactions, as diverse as antigen-antibody and lectin-carbohydrate complexes.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Técnicas Biossensoriais
/
Proteínas
/
Espectrometria de Massas por Ionização por Electrospray
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Mapeamento de Interação de Proteínas
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Anticorpos Imobilizados
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2010
Tipo de documento:
Article