Surface-plasmon-coupled emission: new technology for studying molecular processes.
Explore the source record for details and available documents.
Algorithms↗
Biomedical subjects
Joseph R Lakowicz
Publications and source records attributed to Joseph R Lakowicz.
Directional surface plasmon coupled emission.
Explore the source record for details and available documents.
News↗
Saccharide sensing using gold and silver nanoparticles--a review.
We review new methodologies for glucose sensing from our laboratories based on the specific biological interactions between Con A, dextran-coated gold nanoparticles and glucose, and the interactions between dextran, glucose, and boronic-acid capped silver nanoparticles in solution. Our new approaches promise new tunable glucose sensing platforms. Dextran-coated gold nanoparticles were aggregated with the addition of Con A resulting in increase an in absorbance of nanoparticles at 650 nm, where the post-addition of glucose caused the dissociation of the aggregates and thus a decrease in the absorbance at 650 nm. The interaction of glucose and dextran with boronic acid-capped silver nanoparticles in solution resulted in enhanced luminescence intensity cumulatively due to surface-enhanced fluorescence and the decrease in absorbance at 400 nm, with an increase in absorbance at 640 nm. Lifetime measurements were used to distinguish the contribution from the surface-enhanced fluorescence. TEM was employed to assess the aggregation of nanoparticles.
Algorithms↗
Fluorescence enhancements on silver colloid coated surfaces.
We observed a strong, more than 16-fold, enhancement of Texas Red-labeled BSA fluorescence emission when deposited on silver colloid coated surfaces (SCCS). The same labeled protein deposited on silver island films (SIFs) showed an approximate 8-fold fluorescence enhancement. The lifetimes of Texas Red-BSA fluorescence are significantly shorter on silvered surfaces than on uncoated quartz substrate indicating a strong change in radiative decay rate of the dyes. We also observed a 36-fold increased brightness of overlabeled fluorescein-HSA deposited on silver colloid coated surface. Stronger enhancement observed for overlabeled Fl-HSA protein indicates that presence of silver particles partially decreased self-quenching. Our results indicate that surfaces coated with silver colloids are valuable substrates for metal-enhanced fluorescence.
Algorithms↗
Advances in surface-enhanced fluorescence.
We report recent achievements in metal-enhanced fluorescence from our laboratory. Several fluorophore systems have been studied on metal particle-coated surfaces and in colloid suspensions. In particular, we describe a distance dependent enhancement on silver island films (SIFs), release of self-quenching of fluorescence near silver particles, and the applications of fluorescence enhancement near metalized surfaces to bioassays. We discuss a number of methods for various shaped silver particle deposition on surfaces.
Algorithms↗
Ophthalmic glucose monitoring using disposable contact lenses--a review.
We have developed a range of disposable and colorless tear glucose sensing contact lenses, using off-the-shelf lenses embedded with new water soluble, highly fluorescent and glucose sensitive boronic acid containing fluorophores. The new lenses are readily able to track tear glucose levels and therefore blood glucose levels, which are ideally suited for potential use by diabetics. The fluorescence responses from the lenses can be monitored using simple excitation and emission detection devices. The novelty of our approach is two fold. Firstly, the notion of sensing extremely low glucose concentrations in tears, which track blood levels, by our contact lens approach, and secondly, the unique compatibility of our new glucose signaling probes with the internal mildly acidic contact lens environment. The new lenses are therefore ideal for the non-invasive and continuous monitoring of tear glucose, with about 15-min response time, and a measured shelf life in excess of 3 months. In this review article, we show that fluorescence based signaling using plastic disposable lenses, which have already been industrially optimized with regard to vision correction and oxygen/analyte permeability etc, may a notable alternative to invasive and random finger pricking, the most widely used glucose monitoring technology by diabetics.
Algorithms↗
Metal-enhanced fluorescence solution-based sensing platform.
In recent years our laboratories have reported the favorable effects for fluorophores placed in close proximity to surface immobilized silver nanostructures. These include; greater quantum yields, reduced lifetimes (increased photostability) and directional emission. However, while these findings are likely to find multifarious applications for surface assays based on enhanced fluorescence detection, a solution based enhanced sensing platform has yet to be realized. In this short, note, we show how SiO2-coated silver colloids, indeed provide for a solution based enhanced fluorescence sensing platform with a 3-5 fold enhancement typically observed.
Colloids↗
Wavelength-ratiometric probes for the selective detection of fluoride based on the 6-aminoquinolinium nucleus and boronic acid moiety.
Herein we report a set of new water-soluble fluorescent probes (N-boronobenzyl-6-aminoquinolinium bromides, BAQBAs) sensitive to aqueous fluoride. These probes shows spectral shifts and intensity changes in the presence of fluoride, in a wavelength ratiometric and colorimetric manner, enabling the detection of fluoride concentrations at visible wavelengths, in the concentration range approximately 1-300 mM. Although the sensing mechanism is different for fluoride as compared to the other halides, we have tested the utility of these probes towards the other halides, and the results reveal that the BAQBAs are in fact potential candidates towards the sensing of the all the halides, but in different concentration ranges. As the probes are based on the boronic acid moiety, which is a well-known fluoride and sugar chelator group, we have investigated the response of sugars (such as glucose and fructose, which are present in biological fluids and foodstuffs) as interferences in fluoride detection using these probes. Interestingly, the BAQBAs show a suppressed sugar response potentially allowing for the predominant fluoride sensitivity. In addition to physiological sugars, we also have assessed the response of aqueous halides as potential interferents, or indeed analytes to be sensed, and show that the new boronic acid containing probes respond well to aqueous fluoride in the presence of a high background of other species, such as in a biological cocktail of 50 mM Glucose, 50 mM aqueous Chloride and 5 mM Fructose.
Aminoquinolines↗
Use of surface plasmon-coupled emission to measure DNA hybridization.
The authors describe a new approach to measuring DNA hybridization based on surface plasmon-coupled emission (SPCE). SPCE is the resonance coupling of excited fluorophores with electron motions in thin metal films, resulting in efficient transfer of energy through the film and radiation into the glass substrate. The authors evaluated the use of SPCE for detection of DNA hybridization. An unlabeled capture biotinylated oligonucleotide was attached near the surface of a thin (50 nm) silver film using streptavidin. The authors then measured the emission intensity of single-stranded Cy5-labeled DNA upon binding to a complementary oligomer attached to a silver film. Hybridization could be detected by an increase in SPCE, which appeared as light radiated into the substrate at a sharply defined angle near 73 degrees from the normal. The largest signals were observed when the excitation angle of incidence equaled the surface plasmon wavelength, but directional emission was also observed without excitation by the surface plasmon evanescent field. The increased intensity is due to proximity to the metal surface, so that hybridization can be detected without a change in the quantum yield of the fluorophore. These results indicate that SPCE can provide highly sensitive real-time measurement of DNA hybridization.
Carbocyanines↗
DNA hybridization using surface plasmon-coupled emission.
We describe a new approach to measuring DNA hybridization using surface plasmon-coupled emission (SPCE). Excited fluorophores are known to couple with surface oscillations of electrons in thin metal films, typically 50 nm thick silver on a glass prism. These surface plasmons then radiate into the glass at a sharply defined angle determined by the emission wavelength and the optical properties of the glass and metal. This radiation has the same spectral profile as the emission spectrum of the fluorophores. We studied the emission due to Cy3-labeled DNA oligomers bound to complementary unlabeled oligomers which were themselves bound to the metal surface. Hybridization resulted in SPCE due to Cy3-DNA into the prism. Directional SPCE was observed whether the sample was illuminated from the sample side or through the glass substrate at the surface plasmon angle for the excitation wavelength. A large fraction of the total potential emission is coupled to the surface plasmons resulting in improved sensitivity. When illuminated through the prism at the surface plasmon angle, the sensitivity is increased due to the enhanced intensity of the resonance evanescent field. It is known that SPCE depends on proximity to the silver surface. As a result, changes in emission intensity are observed due to fluorophore localization even if hybridization does not affect the quantum yield of the fluorophore. The use of SPCE resulted in suppression of interfering emission from a noncomplementary Cy5-DNA oligomers due to weaker coupling of the more distant fluorophores with the surface plasmons. We expect SPCE to have numerous applications to nucleic acid analysis and for the measurement of bioaffinity reactions.
DNA↗
Release of the self-quenching of fluorescence near silver metallic surfaces.
Fluorescein is one of the most widely used fluorescent probes in microscopy, biotechnology, and clinical assays. One difficulty with fluorescein is its self-quenching, which results in decreased intensities with increasing labeling density. In this study we examined human serum albumin (HSA), which contained one to nine covalently linked fluorescein molecules per molecule of HSA. The occurrence of homo resonance energy transfer for labeling ratios greater than 1 were confirmed by decreases in the relative quantum yields, anisotropies, and lifetimes. We found that most of the self-quenching can be partially eliminated by proximity of the labeled protein to metallic silver particles. These results suggest the use of heavily labeled proteins and metallic colloids to obtain ultrabright reagents for use in immunoassays, imaging, and other applications.
Fluorescein-5-isothiocyanate↗
Enhanced emission of highly labeled DNA oligomers near silver metallic surfaces.
Fluorescein is a widely used fluorescent probe in DNA analysis. One difficulty with fluorescein is its self-quenching due to resonance energy transfer between the residues, which results in decreased intensities with increasing labeling density. We examined the emission spectral properties of DNA oligomers labeled with one or five fluorescein residues. The emission intensity of the more highly labeled oligomer was decreased due to self-quenching. The self-quenching was mostly eliminated when this oligomer was held approximately 90 A from the surface of metallic silver particles. The intensities increased 7- and 19-fold for the oligomers with one or five fluoresceins, respectively. The increased intensity did not result in increased photobleaching. These results suggest the use of substrates coated with silver particles for increased sensitivity on DNA arrays or for DNA analysis.
Animals↗
Directional surface plasmon-coupled emission: A new method for high sensitivity detection.
Fluorescence emission is nearly isotropic in space. With typical optical components the collection efficiency is 1% or less. In this preliminary report, we describe a novel approach to transforming the normally isotropic emission into directional emission with a collection efficiency near 50%. This can be accomplished for fluorophores located near a semi-transparent silver film on a glass substrate. The emission couples with the surface plasmon resonance on the silver surface and enters the transparent substrate at a sharply defined angle, the surface plasmon angle for the emission wavelength. We estimate that 40-70% of the total emission enters the substrate at the plasmon angle and can thus be directed towards a detector. Background emission from fluorophores distant from the silver does not couple with the plasmon and is not detected. Different emission wavelengths couple at different angles allowing spectral discrimination without additional optics. Surface plasmon-coupled emission represents a new technology which can be used for high detection efficiency with microfluidic and/or surface-bound assay formats.
Fluorescent Dyes↗
Metallic colloid wavelength-ratiometric scattering sensors.
Gold and silver colloids display strong colors as a result of electron oscillations induced by incident light, which are referred to as the plasmon absorption. This absorption is dependent on colloid-colloid proximity, which has been the basis of absorption assays using colloids. We now describe a new approach to optical sensing using the light scattering properties of colloids. Colloid aggregation was induced by avidin-biotin interactions, which shifted the plasmon absorption to longer wavelengths. We found the spectral shift results in changes in the scattering at different incident wavelengths. By measuring the ratio of scattered intensities at two incident wavelengths, this measurement was made independent of the total colloid concentration. The high scattering efficiency of the colloids resulted in intensities equivalent to fluorescence when normalized by the optical density of the fluorophore and colloid. This approach can be used in a wide variety of assay formats, including those commonly used with fluorescence detection.
Affinity Labels↗
DNA hybridization assays using metal-enhanced fluorescence.
We describe a new approach to DNA hybridization assays using metal-enhanced fluorescence. Thiolated oligonucleotides were bound to silver particles on a glass substrate. Addition of a complementary fluorescein-labeled oligonucleotide resulted in a dramatic time-dependent 12-fold increase in fluorescence intensity during hybridization. Proximity to silver particles resulted in a decreased fluorescence lifetime. This effect is thought to be the result of enhanced fluorescence from fluorescein near metallic silver particles. Hybridization could thus be measured from the decay kinetics of the emission, which can be measured independently from the emission intensity. These results suggest the use of silver particles as a general approach to measure DNA hybridization as a method to increase the sensitivity of DNA detection.
Base Sequence↗
Fluorescence spectral properties of cyanine dye-labeled DNA oligomers on surfaces coated with silver particles.
We examined the fluorescence spectral properties of DNA oligomers, labeled with Cy3 or Cy5, when bound to quartz surfaces coated with metallic silver particles. Prior to binding of labeled DNA the surfaces were treated with polylysine or 3-aminopropyl triethoxysilane or were coated with avidin for binding of biotinylated oligomers. The fluorescence intensities were increased an average of 8-fold on these surfaces. Despite the increased emission intensity, the photostability of the labeled DNA was the same or higher on the silver-coated surfaces than on the uncoated slides. The time-integrated intensities, that is the area under the intensity plots with continuous illumination, increased an average of 6-fold. In all cases the lifetimes were dramatically shortened on the silver particles, indicating an over 100-fold increase in the radiative decay rates. These results suggest the use of substrates containing silver particles for increased sensitivity of DNA detection on DNA arrays.
Avidin↗
Increased resonance energy transfer between fluorophores bound to DNA in proximity to metallic silver particles.
We examined the effects of metallic silver particles on resonance energy transfer (RET) between fluorophores covalently bound to DNA. A coumarin donor and a Cy3 acceptor were positioned at opposite ends of a 23-bp double helical DNA oligomer. In the absence of silver particles the extent of RET is near 9%, consistent with a Forster distance R(0) near 50 A and a donor to acceptor distance near 75 A. The transfer efficiency increased when the solution of AMCA-DNA-Cy3 was placed between two quartz plates coated with silver island films to near 64%, as determined by both steady-state and time-resolved measurements. The apparent R(0) in the presence of silver island films increases to about 110 A. These values of the transfer efficiency and R(0) represent weighted averages for donor-acceptor pairs near and distant from the metallic surfaces, so that the values at an optimal distance are likely to be larger. The increased energy transfer is observed only between two sandwiched silvered slides. When we replaced one silvered slide with a quartz plate the effect vanished. Also, the increased energy transfer was not observed for silvered slides separated more than a few micrometers. These results suggest the use of metal-enhanced RET in PCR, hybridization, and other DNA assays, and the possibility of controlling energy transfer by the distance between silver surfaces.
Base Sequence↗
Effects of fluorophore-to-silver distance on the emission of cyanine-dye-labeled oligonucleotides.
We examined the fluorescence spectral properties of Cy3- and Cy5-labeled oligonucleotides at various distances from the surface of silver island films. The distance to the surface was controlled by alternating layers of biotinylated bovine serum albumin (BSA) and avidin, followed by binding of a biotinylated oligonucleotide. The maximum enhancement of fluorescence near a factor of 12 was observed for the first BSA-avidin layer, with the enhancement decreasing to 2-fold for six layers. The minimum lifetimes were observed for the first BSA-avidin layer, and were about 25-fold shorter than on quartz slides without silver, with the lifetimes being about 2-fold shorter for six BSA-avidin layers. These results suggest that maximum fluorescence enhancements occur about 90A from the silver surface, a distance readily obtained by one or two layers of proteins.
Animals↗