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- 2011 (28) (entfernen)
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- Fluorescence (11)
- Nanoparticles (5)
- NIR (4)
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Eingeladener Vortrag
- nein (6)
Recent developments of fluorescent probes beyond approved indocyanine green (ICG) – itself increasingly spreading into new imaging applications like lymphatic mapping, arthritis imaging and tumor surgery – exploit various photophysical and biochemical mechanisms to monitor molecular events with higher specificity and accuracy. Emphasizing nanoparticulate formulations, targeted conjugates, activatable probes, probes with a sensor function and multimodality probes, this review discusses advantages and limitations of each type of probe, thereby critically assessing the desired translation into the clinic.
Labeling of RGD peptides with near-infrared fluorophores yields optical probes for noninvasive imaging of tumors overexpressing ανβ3 integrins. An important prerequisite for optimum detection sensitivity in vivo is strongly absorbing and highly emissive probes with a known fluorescence lifetime. The RGD-Cy5.5 optical probe was derived by coupling Cy5.5 to a cyclic arginine–glycine–aspartic acid–D-phenylalanine–lysine (RGDfK) peptide via an aminohexanoic acid spacer. Spectroscopic properties of the probe were studied in different matrices in comparison to Cy5.5. For in vivo imaging, human glioblastoma cells were subcutaneously implanted into nude mice, and in vivo fluorescence intensity and lifetime were measured. The fluorescence quantum yield and lifetime of Cy5.5 were found to be barely affected on RGD conjugation but dramatically changed in the presence of proteins. By time domain fluorescence imaging, we demonstrated specific binding of RGD-Cy5.5 to glioblastoma xenografts in nude mice. Discrimination of unspecific fluorescence by lifetime-gated analysis further enhanced the detection sensitivity of RGD-Cy5.5-derived signals. We characterized RGD-Cy5.5 as a strongly emissive and stable probe adequate for selective targeting of ανβ3 integrins. The specificity and thus the overall detection sensitivity in vivo were optimized with lifetime gating, based on the previous determination of the probés fluorescence lifetime under application-relevant conditions.
We present the synthesis and characterization of multifunctional fluorophore-labeled poly(organosiloxane) nanoparticles with core-shell architecture, where the fluorescent dye is incorporated into the core. Grafting of heterobifunctional poly(ethylene oxide) (PEO) onto the particle surface leads to water-soluble biocompatible nanoparticles. Two different strategies have been used for the synthesis: The encapsulation of dye-labeled monomers during the polycondensation with additional PEO coating and subsequent dye labeling by covalent attachment of the fluorescent dye rhodamine B to the (chloromethylphenyl)siloxane groups in the core after polymerization and grafting of PEO onto the surface. Comparison of the fluorescence quantum yields of the nanoparticles before and after PEO coating show a decrease in quantum yield after PEO coating.
Polystyrene nanoparticles (PS-NPs) were doped with an oxygen-sensitive near-infrared (NIR)-emissive palladium meso-tetraphenylporphyrin and an inert reference dye which are both excitable at 635 nm. The nanosensors were characterized with special emphasis on fundamental parameters such as absolute photoluminescence quantum yield and fluorescence lifetime. The PS-NPs were employed for ratiometric dual-wavelength and lifetime-based photoluminescent oxygen sensing. They were efficiently taken up by cultured murine alveolar macrophages, yielding a characteristic and reversible change in ratiometric response with decreasing oxygen concentration. This correlated with the cellular hypoxic status verified by analysis of hypoxia inducible factor-1α (HIF-1α) accumulation. In addition, the surface of PS-NPs was functionalized with polyethylene glycol (PEG) and the monoclonal antibody herceptin, and their binding to HER2/neu-overexpressing tumor cells was confirmed in vitro. First experiments with tumor-bearing mouse revealed a distinctive ratiometric response within the tumor upon hypoxic condition induced by animal sacrifice. These results demonstrate the potential of these referenced NIR nanosensors for in vitro and in vivo imaging that present a new generation of optical probes for oncology.
Dye-biomolecule conjugates and NIR-fluorescent particles for targeting of disease-related biomarkers
(2011)
Indispensable for fluorescence imaging are highly specific and sensitive molecular probes that absorb and emit in the near infrared (NIR) spectral region and respond to or target molecular species or processes. Here, we present approaches to targeted fluorescent probes for in vivo imaging in the intensity and lifetime domain exploiting NIR dyes. Screening schemes for the fast identification of suitable fluorophores are derived and design criteria for highly emissive optical probes. In addition, as a signal amplification strategy that enables also the use of hydrophobic NIR fluorophores as fluorescent reporters, first steps towards versatile strategies for the preparation of NIR-fluorescent polymeric particles are presented that can be utilized also for the design of targeted and analyte-responsive probes.
To assess the suitability of asymmetric cyanine dyes for in vivo fluoro-optical molecular imaging, a comprehensive study on the influence of the number of negatively charged sulfonate groups governing the hydrophilicity of the DY-67x family of asymmetric cyanines was performed. Special attention was devoted to the plasma protein binding capacity and related pharmacokinetic properties. Four members of the DY-67x cyanine family composed of the same main chromophore, but substituted with a sequentially increasing number of sulfonate groups (n = 1−4; DY-675, DY-676, DY-677, DY-678, respectively), were incubated with plasma proteins dissolved in phosphate-buffered saline. Protein binding was assessed by absorption spectroscopy, gel electrophoresis, ultrafiltration, and dialysis. Distribution of dye in organs was studied by intraveneous injection of 62 nmol dye/kg body weight into mice (n = 12; up to 180 minutes postinjection) using whole-body near-infrared fluorescence imaging. Spectroscopic studies, gel electrophoresis, and dialysis demonstrated reduced protein binding with increasing number of sulfonate groups. The bovine serum albumin binding constant of the most hydrophobic dye, DY-675, is 18 times higher than that of the most hydrophilic fluorophore, DY-678. In vivo biodistribution analysis underlined a considerable influence of dye hydrophilicity on biodistribution and excretion pathways, with the more hydrophobic dyes, DY-675 and DY-676, accumulating in the liver, followed by strong fluorescence signals in bile and gut owing to accumulation in feces and comparatively hydrophilic DY-678-COOH accumulating in the bladder. Our results demonstrate the possibility of selectively controlling dye-protein interactions and, thus, biodistribution and excretion pathways via proper choice of the fluorophore's substitution pattern. This underlines the importance of structure-property relationships for fluorescent labels. Moreover, our data could provide the basis for the rationalization of future contrast agent developments.