Human alpha-fetal protein immunoassay using fluorescence suppression with fluorescent-bead/antibody conjugate and enzymatic reaction.
Biosens Bioelectron
; 71: 115-120, 2015 Sep 15.
Article
em En
| MEDLINE
| ID: mdl-25897880
ABSTRACT
The aim of the study was to develop a simple and rapid immunoassay using fluorescent microbeads and enzyme-substrate reactions to measure alpha-fetal protein (AFP) concentrations. We demonstrated the functionality of the fluorescent immunosensor using antibody-conjugated fluorescent latex beads (AB-FLBs) and horseradish peroxidase (HRP) to catalyze a reaction, where the products would precipitate and suppress the fluorescence of AB-FLBs. First, the AB-FLBs were incubated with antigen, biotinylated antibodies (bABs), and streptavidin-HRP (SAv-HRP) to form a sandwich-type immunoreaction. The mixture was then filtered through a membrane to concentrate the beads on a small area. After washing to remove unbound bABs and SAv-HRP, a chromogenic HRP substrate and H2O2 were added to form precipitates on the FLB surface. The suppression of the fluorescence was measured with a fluorescent image analyzer system. Under optimized conditions, AFP could be measured at concentrations as low as 1 pg mL(-1) with a dynamic range up to 100 ng mL(-1).
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Alfa-Fetoproteínas
/
Imunofluorescência
/
Imunoconjugados
Tipo de estudo:
Evaluation_studies
Limite:
Humans
Idioma:
En
Revista:
Biosens Bioelectron
Assunto da revista:
BIOTECNOLOGIA
Ano de publicação:
2015
Tipo de documento:
Article