Time-resolved FRET -based approach for antibody detection - a new serodiagnostic concept.
PLoS One
; 8(5): e62739, 2013.
Article
in En
| MEDLINE
| ID: mdl-23667515
ABSTRACT
Förster resonance energy transfer (FRET) is a phenomenon widely utilized in biomedical research of macromolecular interactions. In FRET energy is transferred between two fluorophores, the donor and the acceptor. Herein we describe a novel approach utilizing time-resolved FRET (TR-FRET) for the detection of antibodies not only in a solution-phase homogenous assay but also in single- and two-step solid-phase assays. Our method is based on the principle that the Y-shaped immunoglobulin G molecule is able to simultaneously bind two identical antigen molecules. Hence, if a specific IgG is mixed with donor- and acceptor-labeled antigens, the binding of antigens can be measured by TR-FRET. Using donor- and acceptor-labeled streptavidins (SAs) in conjunction with a polyclonal and a monoclonal anti-SA antibody we demonstrate that this approach is fully functional. In addition we characterize the immune complexes responsible for the TR-FRET signal using density gradient ultracentrifugation and solid-phase immunoassays. The homogenous TR-FRET assay described provides a rapid and robust tool for antibody detection, with a wide potential in medical diagnostics.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Serologic Tests
/
Fluorescence Resonance Energy Transfer
/
Antibodies
Type of study:
Diagnostic_studies
Limits:
Animals
Language:
En
Journal:
PLoS One
Journal subject:
CIENCIA
/
MEDICINA
Year:
2013
Document type:
Article
Affiliation country:
Finland