FRET-based homogeneous immunoassay on a nanoparticle-based photonic crystal

The potential of fluorescence resonance energy transfer (FRET) in a photonic crystal (PC) nanostructured array to enhance the speed and sensitivity of a protein-based immunoassay was tested. Forty-nanometer carboxylated particles conjugated with donor-labeled capture antibodies were trapped by electrophoresis and used as a FRET energy donor. The PC array was able to enhance fluorescent excitation and emission by phase matching. To provide a proof of concept for this FRET-based homogeneous assay on a PC chip, an immunoassay was tested with a simple immunoglobulin G (IgG)-based reaction. A standard curve was generated by testing two different antibody reaction times: 20 min. and 1 min. The results were compared directly to those obtained from a FRET assay that used a modern, high-sensitivity plate reader with a 96-well plate and a reaction time of 1 h. The rabbit-IgG detection limits of the FRET-based homogeneous assay on the PC were 0.001 and 0.1 μg/mL for incubation times of 20 and 1 min, respectively; the sensitivities were 10(3) and 10 times better than the 96-well plate reader, respectively. Thus, FRET on a PC immunoplatform was shown to be a facile, effective, rapid, and sensitive detection technology.