Fluorescence resonance energy transfer in doubly-quantum dot labeled IgG system

The mouse immunoglobulin G (mouse IgG) as a kind of bio-molecule was labeled with two different luminescent colloidal semiconductor quantum dots (QDs), green-emitting CdTe quantum dots and red-emitting CdTe quantum dots in this work. As a result of the fluorescence resonance energy transfer (FRET) between the two different sizes nanoparticles with mouse IgG as the binding bridge, a significant enhancement of the emission of the red-emitting CdTe quantum dots and the corresponding quenching of the emission of green-emitting CdTe quantum dots were observed. The relationship between the concentration of the mouse immunoglobulin G and the fluorescence intensity ratio (I_a/I_d) of acceptors and donors was studied also. Under optimal conditions, the calibration graph is linear over the range of 0.1–20.0 mg/L mouse IgG.     

Spectral characterization of a newly synthesized fluorescent semicarbazone derivative and its usage as a selective fiber optic sensor for copper(II)

In this work photoluminescent properties of highly Cu(2+) selective organic fluoroionophore, semicarbazone derivative; bis(naphtho[2,1-b]furan-2-yl)methanone semicarbazone (BNF) was investigated in different solvents (dichloromethane, tetrahydrofuran, toluene and ethanol) and in polymer matrices of polyvinylchloride (PVC) and ethyl cellulose (EC) by absorption and emission spectrometry. The BNF derivative displayed enhanced fluorescence emission quantum yield, Q(f)=6.1 x 10(-2) and molar extinction coefficient, epsilon=29,000+/-65 cm(-1)M(-1) in immobilized PVC matrix, compared to 2.6 x 10(-3) and 24,573+/-115 in ethanol solution. The offered sensor exhibited remarkable fluorescence intensity quenching upon exposure to Cu(2+) ions at pH 4.0 in the concentration range of 1.0 x 10(-9) to 3.0 x 10(-4)M [Cu(2+)] while the effects of the responding ions (Ca(2+), Hg(+), Pb(2+), Al(3+), Cr(3+), Mn(2+), Mg(2+), Sn(2+), Cd(2+), Co(2+) and Ni(2+)) were less pronounced.     

Upconversion nanoparticle-based fluorescence resonance energy transfer assay for Cr(III) ions in urine

A novel assay of chromium(III) ion based on upconversion fluorescence resonance energy transfer was designed and established. Lysine-capped NaYF₄:Yb/Er upconversion nanoparticles (UCNPs) and dimercaptosuccinic acid-capped gold nanoparticles (AuNPs) were used as the energy donor and acceptor, respectively. They were bound together via electrostatic interaction, resulting in the quenching of the fluorescence of UCNPs by AuNPs. Chromium(III) ions can specifically and strongly interact with dimercaptosuccinic acid that was modified on the surface of AuNPs, leading to the separation of AuNPs from UCNPs and the recovery of fluorescence of UCNPs. The fluorescence recovery of UCNPs showed a good linear response to Cr³⁺ concentration in the range of 2–500 nM with a detection limit of 0.8 nM. This method was further applied to determine the levels of Cr³⁺ in urine. Compared with other fluorescence methods, current method displayed very high sensitivity and signal-to-noise ratio because of the excitation of near-infrared that can eliminate autofluorescence, providing a promising examination of biological samples for the diagnostic purposes.     

Polyethyleneimine-templated copper nanoclusters via ascorbic acid reduction approach as ferric ion sensor

In this report we reported a facile one-pot method for synthesis of water-soluble and stable fluorescent CuNCs at room temperature, in which branched polyethyleneimine (BPEI) served as capping scaffold and ascorbic acid as reducing agent. The prepared BPEI-CuNCs exhibited excellent properties such as good water-solubility, photostability and high stability toward high ionic strength. Based on the electron transfer induced fluorescence quenching mechanism, this fluorescence probe was used for the sensitive and selective determination of ferric ions (Fe³⁺) in aqueous solution. The limit of detection was 340 nM in the linear range of 0.5–1000 μM, which was lower than the maximum level of Fe³⁺ permitted in drinking water by the U.S. Environmental Protection Agency. The method was successfully applied to the detection of Fe³⁺ in tap water, Yellow River water and human urine samples with the quantitative spike recoveries ranging from 95.3% to 112.0%.     

Fluorescence resonance energy transfer (FRET) in chemistry and biology: Non-F?rster distance dependence of the FRET rate

Fluorescence resonance energy transfer (FRET) is a popular tool to study equilibrium and dynamical properties of polymers and biopolymers in condensed phases and is now widely used in conjunction with single molecule spectroscopy. In the data analysis, one usually employs the F?rster expression which predicts (l/R ⁶) distance dependence of the energy transfer rate. However, critical analysis shows that this e_xpression can be of rather limited validity in many cases. We demonstrate this by explicitly considering a donor-acceptor system, polyfluorene (PF₆)-tetraphenylporphyrin (TPP), where the size of both donor and acceptor is comparable to the distance separating them. In such cases, one may expect much weaker distance (as l/R ² or even weaker) dependence. We have also considered the case of energy transfer from a dye to a nanoparticle. Here we find l/R ⁴ distance dependence at large separations, completely different from F?rster. We also discuss recent application of FRET to study polymer conformational dynamics. Dedicated to Prof J Gopalakrishnan on his 62nd birthday.     

A novel fluorescent sensor for Sn4+ detection: Dark resonance energy transfer from silole to rhodamine

A novel dark resonance energy transfer (DRET) off–on cassette SR1 was constructed by coupling a silole donor with a rhodamine acceptor. Due to the intramolecular rotations of the phenyl rings, the silole fluorophore served as a dark donor in solution state and fluorescence leakage from the donor emission could be avoided. Binding with Sn⁴⁺ ion induced the ring‐opening of the rhodamine acceptor, thus increase the overlapping between the emission spectra of the donor and absorption spectra of the acceptor. DRET was turned on and energy was transferred from the silole donor to the rhodamine acceptor. Emission from the rhodamine acceptor was achieved with a large Stokes shift up to 198 nm. The sensor showed good sensitivity and selectivity towards Sn⁴⁺ to other metal ions in methanol aqueous solution through the formation of a 1:1 complex between SR1 and Sn⁴⁺. This research provides a new approach for the development of rhodamine‐based sensors towards metal ions with large Stokes shifts.     

Homogeneous non-competitive bioaffinity assay based on fluorescence resonance energy transfer

A homogeneous non-competitive assay principle for measurement of small analytes based on quenching of fluorescence is described. Fluorescence resonance energy transfer (FRET) occurs between the donor, intrinsically fluorescent europium(III)-chelate conjugated to streptavidin, and the acceptor, quencher dye conjugated to biotin derivative when the biotin-quencher is bound to Eu-streptavidin. Fluorescence can be measured only from those streptavidins that are bound to biotin of the sample, while the fluorescence of the streptavidins that are not occupied by biotin are quenched by quencher-biotin conjugates. The quenching efficiencies of the non-fluorescent quencher dyes were over 95% and one dye molecule was able to quench the fluorescence of more than one europium(III)-chelate. This, however, together with the quadrovalent nature of streptavidin limited the measurable range of the assay to 0.2-2 nmol L⁻¹. In this study we demonstrated that FRET could be used to design a non-competitive homogeneous assay for a small analyte resulting in equal performance with competitive heterogeneous assay.     

Fluorescence resonance energy transfer from tryptophan in human serum albumin to a bioactive indoloquinolizine system

The interaction between a bioactive molecule, 3-acetyl-4-oxo-6,7-dihydro-12H indolo-[2,3-a] quinolizine (AODIQ), with human serum albumin (HSA) has been studied using steady-state absorption and fluorescence techniques. A 1:1 complex formation has been established and the binding constant (K) and free energy change for the process have been reported. The AODIQ-HSA complex results in fluorescence resonance energy transfer (FRET) from the tryptophan moiety of HSA to the probe. The critical energy-transfer distance (R ₀) for FRET and the Stern-Volmer constant (K _sv) for the fluorescence quenching of the donor in the presence of the acceptor have been determined. Importantly, K _SV has been shown to be equal to the binding constant itself, implying that the fluorescence quenching arises only from the FRET process. The study suggests that the donor and the acceptor are bound to the same protein at different locations but within the quenching distance.     

Fluorescence energy transfer probes based on the guanine quadruplex formation for the fluorometric detection of potassium ion

Dual-labeled oligonucleotide derivative, FAT-0, carrying 6-carboxyfluorescein (FAM) and 6-carboxy-tetramethylrhodamine (TAMRA) labels at 5′- and 3′-termini of thrombin-binding aptamer (TBA) sequence 5′-GGTTGGTGTGGTTGG-3′ and its derivatives, FAT-n (n = 3, 5, and 7) were designed and synthesized. FAT-n derivatives contained a T_mA spacer (m = 2, 4, and 6, respectively) at 5′-end of TBA sequence. The probes were developed to estimate the spacer effect on FRET efficiency and to identify the best probe for sensing of K⁺. Circular dichroism (CD), UV-vis absorption, and fluorescence studies revealed that all FAT-n probes could form the intramolecular tetraplex structures after binding K⁺. Association constants of particular K⁺/FAT-n complexes were determined using different experimental approaches. Suitability of particular probes for sensitive monitoring of K⁺ in intra- and extracellular conditions was examined and discussed. Calibration graphs of fluorescence ratio were linear in the K⁺ concentration range of 2-10 mM for extracellular conditions showing sensitivity of 1.2% mM⁻¹ K⁺ and for intracellular conditions in the range of 100-200 mM with sensitivity of 0.49% mM⁻¹ K⁺.     

Lineal DNA logic gate for microRNA diagnostics with strand displacement and fluorescence resonance energy transfer

Designing molecular logic gates to operate programmably for molecular diagnostics in molecular computing still remains challenging. Here, we designed a novel linear DNA logic gates for microRNA analysis based on strand displacement and fluorescence resonance energy transfer (FRET). Two labeled strands closed each other produce to FRET through hybridization with a complementary strand to form a basic work unit of logic gate. Two indicators of heart failure (microRNA-195 and microRNA-21) were selected as the logic inputs and the fluorescence mode was used as the logic output. We have demonstrated that the molecular logic gate mechanism worked well with the construction of YES and AND gates.     

Synthesis and characterization of anthracene-clustering dendrimers: observation of fluorescence resonance energy transfer in the multichromophoric system

A series of anthracene-clustering dendrimers bearing various aliphatic substituents at the terminal positions were synthesized using a direct coupling strategy. A remarkable effect of the side chains was imparted to chemical properties of the dendrimers such as drastically increased solubility. Although the multibranched anthracene arrays in the dendritic architectures exhibited no cooperativity in terms of the absorption feature and behaved as single chromophoric systems, investigations focusing on fluorescence properties revealed that a type of cooperativity was present as expressed in the reduced quantum yields of fluorescence. An alternative approach utilizing time-resolved fluorescence decay measurements clearly demonstrated that the most reasonable mechanism of the cooperative action should involve two discernible channels of intramolecular fluorescence resonance energy transfer (FRET) occurring from one chromophore to the others within and across junctions of the branching units.     

Fluorescence resonance energy transfer by quencher adsorption into hydrogels containing fluorophores

We synthesized anionic hydrogels containing fluorophores and investigated the adsorption of a cationic quencher having an amino group into hydrogels by fluorescence resonance energy transfer (FRET). FRET from the fluorophore to the quencher in hydrogels was examined by fluorescence intensity and fluorescence decay using a fluorescence spectrophotometer and femtosecond laser spectroscopy. The fluorescence intensity of the fluorophore‐containing hydrogels decreased rapidly with increasing amounts of adsorbed cationic quencher. The fluorescence emission of the fluorophore in the quencher‐adsorbed hydrogels containing fluorophores decayed more rapidly than that of the original hydrogels. The aforementioned result indicates that the fluorescence of the fluorophore‐containing hydrogels is quenched due to FRET from the fluorophore to the quencher as the cationic quenchers can approach the fluorophores in hydrogels by electrostatic interactions. © 2006 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 44: 3245–3252, 2006     

A nanoparticle-supported fluorescence resonance energy transfer system formed via layer-by-layer approach as a ratiometric sensor for mercury ions in water

This article describes the design and preparation of a novel fluorescence resonance energy transfer (FRET)-based ratiometric sensor with the polymer nanoparticle as scaffold for detecting Hg²⁺ in aqueous media. In this study, a fluorescent dye fluorescein isothiocyanate (FITC, served as the donor) and a spirolactam rhodamine derivative (SRHB, served as mercury ion probe) were covalently attached onto polyethylenimine (PEI) and polyacrylic acid (PAA) respectively; and a ratiometric sensing system was then formed through the deposition of the donor- and probe-containing polyelectrolytes onto the negatively charged polymer particles via the layer-by-layer approach. The ratiometric fluorescent signal change of the system is based on the intra-particle fluorescence resonance energy transfer (FRET) process modulated by mercury ions. Under optimized structural and experimental conditions, the particle-based detection system exhibits stable response for Hg²⁺ in aqueous media. More importantly, in this newly developed particle-based detection system formed by LBL approach, varied numbers of the PAA/PEI layers which served as the spacer could be placed between the donor-containing layer and the probe-containing layer, hence the donor–acceptor distance and energy transfer efficiency could be effectively tuned (from ca. 25% to 76%), this approach has well solved the problem for many particle-based FRET systems that the donor–acceptor distance cannot be precisely controlled. Also, it is found that the ratiometric sensor is applicable in a pH range of 4.6–7.3 in water with the detection limit of 200 nM. This approach may provide a new strategy for ratiometric detection of analytes in some environmental and biological applications.     

Fluorescence sensing based on cation-induced conformational switching: copper-selective modulation of the photoinduced intramolecular charge transfer of a donor-acceptor biphenyl fluorophore

The fluorescence emission energy of donor-acceptor substituted biphenyls is highly sensitive towards conformational changes of the interannular twist angle. By integrating 4-dimethylamino-4′-cyano-biphenyl into the ligand backbone of a thioether-rich metal receptor we designed a fluorescence sensor that exhibits high selectivity towards copper. Upon metal binding the ligand undergoes a significant conformational change, which induces a strong hypsochromic shift of the photoinduced charge-transfer emission. Steady-state absorption and fluorescence spectroscopy revealed a high affinity towards Cu(I) with a well-defined 1:1 metal-ligand complex stoichiometry. The nature of the conformational changes upon Cu(I) coordination were analyzed in detail by ¹H NMR and 2D NOESY experiments. The spectroscopic data provide a coherent picture, which is consistent with a Boltzmann ground-state distribution of several rotamers that are locked into a more flattened geometry upon coordination of Cu(I).     

Resonance energy transfer from dibucaine to acriflavine in polystyrene latex dispersions

Resonance energy transfer from dibucaine (DC) to acriflavine (AF) has been investigated both in homogeneous aqueous solutions and in polystyrene latex dispersions. The energy transfer reaction is observed by monitoring fluorescence quenching of DC as well as sensitized emission of AF. It is found that the energy transfer from DC to AF is remarkably enhanced on going from the aqueous homogeneous solution to the latex dispersion. This is mainly attributed to the fact that both the donor and acceptor are effectively adsorbed onto the latex particles, as evidenced by the measurement of adsorption isotherms. From the adsorption experiments, it is also elucidated that electrostatic interaction is significant at low DC concentration, while hydrophobic interaction dominates at the higher concentration especially at higher pH.     

Construction of triblock copolymer-gold nanorod composites for fluorescence resonance energy transfer via pH-sensitive allosteric

To explore the effects of microenvironmental adjustments on fluorescence, a pH-sensitive nanocomposite system based on fluorescence resonance energy transfer (FRET) was constructed. The model system included a modified triblock copolymer (polyhistidine-b-polyethylene glycol-b-polycaprolactone) and gold nanoparticles. A near-infrared dye was used as the donor, and spectrally matched gold nanorods, attached after C-terminus modification with α-lipoic acid, were used as the receptor to realize control of the FRET effect over the fluorescence intensity for two polymer configurational changes (i.e., “folded” and “stretched” states) in response to pH. After synthesis and characterization, we investigated the self-assembly behavior of the system. Analysis by quartz crystal microbalance revealed the pH sensitivity of the polymer, which exhibited “folding” and “stretching” states with changes in pH, providing a structural basis for the FRET effect. Fluorescence spectrophotometry investigations also revealed the regulatory impact of the assembled system on fluorescence.     

A competitive displacement assay with quantum dots as fluorescence resonance energy transfer donors

The unique optoelectronic properties of semiconductor quantum dots (QDs) make them well-suited as fluorescent bioprobes for use in various biological applications. Modification of CdSe/ZnS QDs with biologically relevant molecules provides for multipotent probes that can be used for cellular labeling, bioassays, and localized optical interrogation by means of fluorescence resonance energy transfer (FRET). Herein, we demonstrate the use of red-emitting streptavidin-coated QDs (QD₆₀₅) as donors in FRET to introduce a competitive displacement-based assay for the detection of oligonucleotides. Various QD–DNA bioconjugates featuring 25-mer probe sequences diagnostic of Hsp23 were prepared. The single-stranded oligonucleotide probes were hybridized to dye-labeled (Alexa Fluor 647) reporter sequences, which were provided for a FRET-sensitized emission signal due to proximity of the QD and dye. The dye-labeled sequence was designed to be partially complementary and include base-pair mismatches to facilitate displacement by a more energetically favorable, fully complementary recognition motif embedded within a 98-mer displacer sequence. Overall, this study demonstrates proof-of-concept at the nM level for competitive displacement hybridization assays in vitro by reduction of fluorescence intensity that directly correlates to the presence of oligonucleotides of interest. This work demonstrates an analytical method that could potentially be implemented for monitoring of intracellular gene expression in the future.     

A fast,highly selective and sensitive dansyl-based fluorescent sensor for copper (II) ions and its imaging application in living cells

Since the copper ions (Cu²⁺) play a fatal role in many foundational physiological processes, it is important to develop a simple, highly sensitive and selective sensor for Cu²⁺ detection in living systems. Herein, an intramolecular charge transfer (ICT) and dansyl-based fluorescent chemosensor 1 was designed, synthesized and characterized for the sensitive and selective quantification of Cu²⁺. It exhibited remarkable fluorescence quenching upon addition of Cu²⁺ over other selected metal ions, attributed to the complex formation between 1 and Cu²⁺ with the association constant 6.7 × 10⁵ M^?1. The sensor 1 showed a fast and linear response towards Cu²⁺ in the concentration range from 0 to 12.5 × 10^?6 mol L^?1 with the detection limit of 2.5 × 10^?7 mol L^?1. This detection could be carried out in a wide pH range of 5.0–14. Furthermore, sensor 1 can be used for detecting Cu²⁺ in living cells.     

A new tridentate fluorescent-colorimetric chemosensor for copper(II) ion

A new tri-imidazolium salt, tris(4-(3-(2-((8-hydroxy-9,10-dioxo-9′,10′-dihydroanthracen-1-yl)oxy)ethyl)-1H-imidazole-3′-ium-1′-yl)phenyl)amine hexafluorophosphate was prepared and characterized. Particularly, the recognition performance of the tri-imidazolium salt for cations was investigated through fluorescence and ultraviolet titrations, MS, IR spectra and ¹H NMR titrations. The results indicated that the tri-imidazolium salt can distinguish effectively copper(II) ion from other cations by the changes of spectroscopy and colour (from yellow to orange under sunlight). Furthermore, the tri-imidazolium salt was also used in detecting Cu²⁺ through employing smartphone with the computed detection limit down to 0.51 μM.     

Study on the fluorescence resonance energy transfer between CdTe QDs and butyl-rhodamine B in the presence of CTMAB and its application on the detection of Hg(II)

The thioglycolic acid-functionalized CdTe quantum dots (QDs) were synthesized in aqueous solution using safe and low-cost inorganic salts as precursors. Fluorescence resonance energy transfer (FRET) system was constructed between CdTe QDs (donor) and butyl-rhodamine B (BRB) (acceptor) in the presence of cetyltrimethylammonium bromide (CTMAB). CTMAB micelles formed in water reduced the distance between the donor and the acceptor significantly and thus improved the FRET efficiency, which resulted in an obvious fluorescence enhancement of the acceptor. Several factors which impacted the fluorescence spectra of the FRET system were studied. The energy transfer efficiency (E) and the distance (r) between CdTe and BRB were obtained. The feasibility of the prepared FRET system as fluorescence probe for detecting Hg(II) in aqueous solution was demonstrated. At pH 6.60, a linear relationship could be established between the quenched fluorescence intensity of BRB and the concentration of Hg(II) in the range of 0.0625-2.5mumolL(-1). The limit of detection was 20.3nmolL(-1). The developed method was proved to be sensitive and repeatable to detect Hg(II) in a wide range in aqueous solutions.