Europium nanoparticles and time-resolved fluorescence for ultrasensitive detection of prostate-specific antigen.
BACKGROUND
Nanoparticle-based detection technologies have the potential to improve detection sensitivity in miniature as well as in conventional biochemical assays. We introduce a detection technology that relies on the use of europium(III) nanoparticles and time-resolved fluorometry to improve the detection limit of biochemical assays and to visualize individual molecules in a microtiter plate format.


METHODS
Streptavidin was covalently coated on 107-nm nanoparticles containing >30 000 europium molecules entrapped with beta-diketones. In a model assay system, these nanoparticles were used to trace biotinylated prostate-specific antigen (PSA) in a microtiter plate format.


RESULTS
The detection limit (mean + 3 SD of the zero calibrator) of biotinylated PSA was 0.38 ng/L, corresponding to 10 fmol/L or 60 zeptomoles (60 x 10(-21) moles) of PSA. Moreover, single nanoparticles, representing individual PSA molecules, were visualized in the same microtiter wells with a time-resolved fluorescence microscope using a x10 objective. Single nanoparticles, possessing high specific activity, were also detected in solution by a standard time-resolved plate fluorometer.


CONCLUSIONS
The universal streptavidin-coated europium(III) nanoparticle label is suitable for detection of any biotinylated molecule either in solution or on a solid phase. The europium(III) nanoparticle labeling technology is applicable to many areas of modern biochemical analysis, such as immunochemical and multianalyte DNA-chip assays as well as histo- and cytochemistry to improve detection sensitivities.
Department of Biotechnology, University of Turku, Tykistökatu 6, FIN-20520 Turku, Finland. harri.harma@utu.fi
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Härmä
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Soukka
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Lövgren
D019921:Biotinylation; D002614:Chelating Agents; D005063:Europium; D005456:Fluorescent Dyes; D015200:Fluoroimmunoassay; D007089:Image Enhancement; D007202:Indicators and Reagents; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D010316:Particle Size; D017430:Prostate-Specific Antigen / Q000032:analysis*; D019809:Streptavidin
D002614:Chelating Agents; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D005063:Europium; D019809:Streptavidin; D017430:Prostate-Specific Antigen