Fluorescence quenching competitive immunoassay in micro droplets.
A fluorescence quenching competitive immunoassay in micro droplets was applied to the sensitive detection of the pyrethroid insecticide, esfenvalerate. Laser induced fluorescence from rhodamine dye was used as a marker. The competitive immunoreaction was performed in micro droplets generated by a vibrating orifice aerosol generator system with a 10-microm diameter orifice. Fluorescence that was emitted from the droplets was detected by a 1/8 m imaging spectrograph with a 512 x 512 thermoelectrically cooled, charged-coupled device camera. The conjugate of esfenvalerate with rhodamine exhibited similar fluorescence to that of pure rhodamine 6G. When anti-esfenvalerate antibodies were added to the droplets, the fluorescence decreased. The reduction in emission was due to a strong quenching effect that arises from the interaction between the protein and rhodamine molecules following the antigen-antibody reaction. When a sample of esfenvalerate was added to the droplets, the release of the conjugated rhodamine from the antigen-antibody complex allowed the fluorescence signal to recover. An assay in a picoliter droplet sample was shown to enable detection down to approximately 0.1 nM. A very small mass of analyte could be detected with this method. A sample of river water was used to gauge the impact of matrix effects and was shown to give rise to negligible interference with the immunoassay.
Feng
Jun
J
Department of Mechanical and Aeronautical Engineering, University of California, Davis, CA 95616, USA.
Shan
Guomin
G
Hammock
Bruce D
BD
Kennedy
Ian M
IM
D000336:Aerosols / Q000032:analysis*; D000906:Antibodies / Q000032:analysis / Q000276:immunology; D004785:Environmental Pollutants / Q000032:analysis; D004867:Equipment Design; D019544:Equipment Failure Analysis; D016231:Fluorescence Polarization Immunoassay / Q000295:instrumentation* / Q000379:methods; D036681:Fluorescence Recovery After Photobleaching / Q000295:instrumentation* / Q000379:methods; D008832:Microchemistry / Q000295:instrumentation* / Q000379:methods*; D009570:Nitriles; D011722:Pyrethrins / Q000032:analysis* / Q000276:immunology; D015203:Reproducibility of Results; D012235:Rhodamines / Q000737:chemistry*; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000295:instrumentation / Q000379:methods
D000336:Aerosols; D000906:Antibodies; D004785:Environmental Pollutants; D009570:Nitriles; D011722:Pyrethrins; D012235:Rhodamines; C017690:fenvalerate