A new europium(III) complex, [4'-(10-methyl-9-anthryl)-2,2':6',2"-terpyridine-6,6"-diyl]bis(methylenenitrilo) tetrakis(acetate)-Eu(3+), was designed and synthesized as a highly sensitive and selective time-gated luminescence probe for singlet oxygen ((1)O2). The new probe is highly water soluble with a large stability constant of approximately 10(21) and a wide pH available range (pH 3-10), and can specifically react with (1)O2 to form its endoperoxide (EP-MTTA-Eu(3+)) with a high reaction rate constant at 10(10) M(-1) s(-1), accompanied by the remarkable increases of luminescence quantum yield from 0.90% to 13.8% and lifetime from 0.80 to 1.29 ms, respectively. The wide applicability of the probe was demonstrated by detection of (1)O2 generated from a MoO(4)(2-)/H(2)O2 system, a photosensitization system of 5,10,15,20-tetrakis(1-methyl-4-pyridinio)porphyrin tetra(p-toluenesulfonate) (TMPyP), and a horseradish peroxidase catalyzed aerobic oxidation system of indole-3-acetic acid (IAA). In addition, it was found that the new probe could be easily transferred into living HeLa cells by incubation with TMPyP. A time-gated luminescence imaging technique that can fully eliminate the short-lived background fluorescence from TMPyP and cell components has been successfully developed for monitoring the time-dependent generation of (1)O2 in living cells.

Novel silica-coated terbium(III) chelate fluorescent nanoparticles have been prepared and characterized as a new type of fluorescence probe for highly sensitive time-resolved fluorescence bioassay. The preparation was carried out in a water-in-oil microemulsion containing a strongly fluorescent Tb(3+) chelate, N,N,N(1),N(1)-[2,6-bis(3'-aminomethyl-1'-pyrazolyl)-phenylpyridine]tetrakis(acetate)-Tb(3+), Triton X-100, hexanol, and cyclohexane by controlling hydrolysis of tetraethyl orthosilicate. The nanoparticles are spherical and uniform in size, 42 +/- 3 nm in diameter, strongly fluorescent, and highly photostable and have enough of a long fluorescence lifetime (1.52 ms) for time-resolved fluorescence measurement. A stable and nontoxic method was developed for the surface modification and protein immobilization of the nanoparticles. As a model of application, the nanoparticle-labeled streptavidin was prepared and used in a sandwich-type time-resolved fluoroimmunoassay of human prostate-specific antigen (PSA) by using a 96-well microtiter plate as the solid-phase carrier. The method gives a detection limit of 7.0 pg/mL for the PSA assay.

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Novel silica-coated luminescent europium (SCLE) nanoparticles have been prepared by copolymerization of tetraethyl orthosilicate and 3-aminopropyl(triethoxy)silane with ammonium hydroxide in a water-in-oil microemulsion containing a luminescent Eu 3+ chelate, Triton X-100, n-octanol, and cyclohexane. The nanoparticles were characterized by transmission electron microscopy, X-ray diffraction, and UV-vis and luminescence spectra. The results show that the nanoparticles are spherical and uniform in size, 50 ( 5 nm in diameter, and have high photostability and long luminescence lifetime (770 µs). Because primary amino groups have been directly introduced to the nanoparticle's surface by using 3-aminopropyl-(triethoxyl)silane in nanoparticle preparation, the surface modification and bioconjugation of the nanoparticles are easier and reproducible. The nanoparticles were used for streptavidin (SA) labeling, and the nanoparticle-labeled streptavidin was successfully used in timeresolved fluoroimmunoassay of human R-fetoprotein. The results reveal that the new nanoparticles are favorable for use as a new type of luminescence probe for highly sensitive time-resolved luminescence bioassay.

Effective imaging of cancer molecular biomarker is critical for accurate cancer diagnosis and prognosis. CLT1 peptide was observed to specifically bind to the fibrin-fibronectin complexes presented in tumor extracellular matrix. In this study, we synthesized and evaluated CLT1 peptidetargeted nanoglobular Gd-DOTA monoamide conjugates for magnetic resonance (MR) imaging of the fibrin-fibronectin complexes in tumor. The targeted nanoglobular contrast agents were prepared by conjugating peptide CLT1 to G2 and G3 nanoglobule (lysine dendrimers with a cubic silsesquioxane core) Gd-DOTA monoamide conjugates via click chemistry. The T 1 relaxivities of peptide targeted G2 and G3 nanoglobules were 7.92 and 8.20 mM −1 s −1 at 3T, respectively. Approximately 2 peptides and 25 Gd-DOTA chelates were conjugated onto the surface of 32 amine groups of G2 nanoglobule, and 3 peptides and 43 Gd-DOTA chelates onto the surface of 64 amine groups of G3 nanoglobule. The peptide-targeted nanoglobular contrast agents showed greater contrast enhancement than the corresponding non-targeted agents in tumor at a dose of 0.03 mmol/ kg in female athymic mice bearing MDA-MB-231 human breast carcinoma xenografts. The targeted MRI contrast agents have a potential for specific cancer molecular imaging with MRI.

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