Development of a Visible-Light-Sensitized Europium Complex for Time-Resolved Fluorometric Application

摘要

The time-resolved luminescence bioassay technique usingnluminescent lanthanide complexes as labels is a highlynsensitive and widely used bioassay method for clinicalndiagnostics and biotechnology. A major drawback of thencurrent technique is that the luminescent lanthanidenlabels require UV excitation (typically less than 360 nm),nwhich can damage living biological systems and is holdingnback further development of time-resolved luminescenceninstruments. Herein we describe two approaches fornpreparing a visible-light-sensitized Eu3+ complex innaqueous media for time-resolved fluorometric applications:na dissociation enhancement aqueous solutionnthat can be excited by visible light for ethylenediaminetetraacetaten(EDTA)-Eu3+ detection and a visible-lightsensitizednwater-soluble Eu3+ complex conjugated bovinenserum albumin (BSA) for biolabeling and timeresolvednluminescence bioimaging. In the first approach,na weakly acidic aqueous solution consisting of 4,4′-nbis(1′′,1′′,1′′,2′′,2′′,3′′,3′′-heptafluoro-4′′,6′′-hexanedion-n6′′-yl)-o-terphenyl (BHHT), 2-(N,N-diethylanilin-4-yl)-n4,6-bis(3,5-dimethylpyrazol-1-yl)-1,3,5-triazine (DPBT),nand Triton X-100 was prepared. This solution showsna strong luminescence enhancement effect fornEDTA-Eu3+ with a wide excitation wavelength rangenfrom UV to visible light (a maximum at 387 nm) and anlong luminescence lifetime (520 μs), to provide a novelndissociation enhancement solution for time-resolvednluminescence detection of EDTA-Eu3+. In the secondnapproach, a ternary Eu3+ complex, 4,4′-bis(1′′,1′′,1′′,2′′,-n2′′,3′′,3′′-heptafluoro-4′′,6′′-hexanedion-6′′-yl)-chlorosulfo-no-terphenyl (BHHCT)-Eu3+-DPBT, was covalentlynbound to BSA to form a water-soluble BSA-BHHCTEu3+-nDPBT conjugate. This biocompatible conjugatenis of the visible-light excitable feature in aqueous medianwith a wide excitation wavelength range from UV tonvisible light (a maximum at 387 nm), a long luminescencenlifetime (460 μs), and a higher quantum yieldn(27%). The conjugate was successfully used for streptavidinn(SA) labeling and time-resolved luminescencenimaging detection of three environmental pathogens,nGiardialamblia,Cryptosporidiummuris,andCryptosporidiumnparvum, in water samples. Our strategy givesna general idea for designing a visible-light-sensitizednEu3+ complex for time-resolved luminescence bioassaynapplications.