Time-resolved Fluorescence Spectroscopy Research Articles

6-(6-Methyl-1,2,4,5-Tetrazine-3-yl)-2,2'-Bipyridine: A N-Donor Ligand for the Separation of Lanthanides(III) and Actinides(III).

Here, we report the synthesis of the 6-(6-methyl-1,2,4,5-tetrazine-3-yl)-2,2'-bipyridine (MTB) ligand that has been developed for lanthanide/actinide separation. A multimethod study of the complexation of MTB with trivalent actinide and lanthanide ions is presented. Single-crystal X-ray diffraction measurements reveal the formation of [Ce(MTB)2(NO3)3], [Pr(MTB)(NO3)3H2O], and [Ln(MTB)(NO3)3MeCN] (Ln = Nd, Sm, Eu, Gd). In addition, the complexation of Cm(III) with MTB in solution was studied by time-resolved laser fluorescence spectroscopy. The results show the formation of [Cm(MTB)1-3]3+ complexes, which occur in two different isomers. Quantum chemical calculations reveal an energy difference between these isomers of 12 kJ mol-1, clarifying the initial observations made by time-resolved laser fluorescence spectroscopy (TRLFS). Furthermore, quantum theory of atoms in molecules (QTAIM) analysis of the Cm(III) and Ln(III) complexes was performed, indicating a stronger covalent contribution in the Cm-N interaction compared to the respective Ln-N interaction. These findings align well with extraction data showing a preferred extraction of Am and Cm over lanthanides (e.g., max. SFAm/Eu = 8.3) at nitric acid concentrations <0.1 mol L-1 HNO3.

Solvent effect on the excited state photophysics of Imiquimod: A DFT/TD-DFT and spectroscopic study

Solvation plays an important role in chemistry and biology. The role of the solvent is crucial in any chemical processes such as tautomerization that controls the structure and function of biomolecules. The current study aims to explore how different solvent medium affects the tautomeric forms of an antitumor drug, Imiquimod (IMQ). We have used several spectroscopical methods, including UV–Vis absorption spectroscopy, steady-state and time-resolved fluorescence spectroscopy, and quantum mechanical (QM) calculations. Our results revealed that solvent, indeed, plays a significant role in the modulation of the photophysics of the drug. IMQ has three emission bands in protic and aprotic solvents and two bands in non-polar medium associated with different forms of the drug. Using time-resolved technique, and comparing with the predicted lifetimes from QM calculations, we succeed to assign these three forms as cation, tautomer and neutral of IMQ with 1.6 ns, 2.0 ns and 4.0 ns lifetime values, respectively. Since IMQ is a nucleobase analogue and one of the most effective medications for skin tumors; these findings about which specie of IMQ is present in a given medium, and beyond, how the medium alters the photophysics of the molecule may provide deeper insights into its structure and function.

An inclusive comparison regarding aggregation of surface active ionic liquid and conventional surfactant with a cationic dye Acridine Red exposed in view of spectroscopic and theoretical study

The investigation was conducted on the interaction of acridine red (AR) with sodium dodecyl sulfate (SDS) and 1-butyl-3-methyl imidazolium octyl sulfate (BMImOS) in both premicellar and post micellar concentrations. The interaction between AR and room temperature ionic liquid (RTIL) 1-butyl-3-methyl imidazolium bromide (BMImBr) was studied to understand the effect of imidazolium cation on micellization. Spectroscopic experiments, such as UV–visible absorption spectroscopy, steady-state fluorescence spectroscopy, time-resolved fluorescence spectroscopy, and dynamic light scattering, have been introduced. Various important parameters like spectral shifts, anisotropy, and aggregation number have been determined spectroscopically. However, a fresh insight into density functional theory calculations has also been provided. The observed results have been explained in terms of the aggregation behavior of the amphiphiles. A prominent redshift accompanied by a change in intensity is observed in the presence of surfactant and SAIL. However, in the case of BMImBr, no such spectral shift has been observed; only a decrease in spectral intensity highlights the quenching ability of the imidazolium cation. Average lifetime also provides some additional information regarding the difference in the formation of aggregates by SDS and BMImOS, which is further established from particle size density obtained from DLS. The salt like role of 1-butyl-3-methyl imidazolium cation also has been logically explained by anisotropy measurement and the determination of aggregation number. Using Gauss View 5.0 and Gaussian 09 package, the energy of optimization of each set of amphiphiles along with dye has been determined to follow the interaction at the molecular level. TDDFT calculations have also been performed to determine the structure of HOMO and LUMO.

Steady State and Time-Resolved Fluorescence Spectroscopy of Cinchonine Dication in Sodium Dodecylsulphate Micellar System.

This paper reports the influence of surface charge of the micelles on to the photophysical properties of a cinchonine dication (C2+) fluorophore in anionic, sodium dodecylsulphate (SDS), surfactant at premicellar, micellar and post-micellar concentrations in aqueous phase at room temperature. The magnitude of edge excitation red shift (EERS) in the fluorescence maximum of C2+ in bulk water solution is 1897cm- 1 whereas, in the case of SDS it is observed to be 1984cm- 1. The fluorescence decay curve of C2+ fits with multi exponential functions in the micellar system. The increase in lifetime of C2+ in SDS has been attributed to the increase in radiative rate due to the incorporation of C2+ at the micelle -water interface. The value of dynamic quenching constant determined is 16.9 M- 1. The location of the probe molecule in micellar systems has been justified by a variety of spectral parameters such as dielectric constant, ET (30), viscosity, EERS, average fluorescence decay time, radiative and non-radiative rate constants. All experimental results suggest that the C2+ molecule binds strongly with the SDS micelles and resides at micellar-water interface. The binding constant (Kb) calculated (3.85 × 105 M- 1) for C2+ in SDS revealed that the electrostatic forces mediate charge probe-micelle association.

Efficiency of Encapsulation of Thioflavin T (ThT) into Polar and Nonpolar Environments of Different Bile Salt Aggregates: A Femtosecond Fluorescence Study.

The binding of Thioflavin T (ThT) with various bile salts, a potential host molecule, has been analyzed by steady-state and time-resolved fluorescence spectroscopy. A comparative study has been executed to investigate the influence of confinement of different bile salts, namely, sodium cholate (NaCh), sodium taurocholate (NaTC), and sodium deoxycholate (NaDC) on binding and excited state torsional motion of ThT molecules. The changes in absorption and emission properties of probe molecules were found to be sensitive to increasing bile salt concentration in aqueous 0.2 (M) NaCl solutions. The photophysics of ThT mainly depends on hydrophobicity, morphology, and size of bile salt aggregates in solution. In the presence of bile salts, the emission intensity and emission lifetime of ThT increase significantly, indicating encapsulation of dye. Moreover, we have also investigated the effect of the ionic strength of the medium by sodium chloride (NaCl) on the spectroscopic properties of ThT in the restricted surroundings of aqueous bile salts. It is observed that the fluorescence lifetime of ThT in bile salts increases significantly in the presence of NaCl. The encapsulation efficiency of ThT in bile salt aggregates has been assessed by iodide (I-) as an external ionic quencher. We found that NaDC aggregates are more efficient in the modulation of photophysical properties of ThT and also provide better protection efficiency to decrease the nonradiative deactivation processes.

Influence of Structural Properties of Oleic Acid-Capped CdSe/ZnS Quantum Dots in the Detection of Hg2+ Ions.

Oleic acid-capped CdSe/ZnS quantum dots (QDs) were used to investigate their photoluminescence (PL) response to Hg2+ ions as a function of the surface properties of QDs. Three distinctly-size CdSe/ZnS QDs were obtained by varying the molar ratio of shell precursors, which were characterized by X-ray diffraction (XRD), high-resolution transmission electron microscopy (HR-TEM), Fourier-Transform infrared (FT-IR), X-ray photoelectron spectroscopy (XPS), absorption spectroscopy, and time-resolved fluorescence spectroscopy. Results revealed the obtention of zinc blende nanocrystals with sizes ranging from 2.7 to 3.2nm (± 0.5) and ZnS thickness between 0.3 and 1.0 monolayer (ML). The variation of the [S]/[Zn] molar ratio introduced chemical species that act as traps, affecting the PL properties differently. Depending on the thickness of the shell and chemical speciation on surface, Hg2+ ions could induce quenching or enhancement of PL. Detection of mercury ions was evaluated in terms of Stern-Volmer equation, where the limit of detection (LOD) for the PL quenching system was 11.2 nM, while for the PL enhancing systems were 8.98 nM and 10.7 nM. Results demonstrate the performance of oleic acid-capped CdSe/ZnS QDs to detect Hg2+ and their capacity to turn the PL on/off depending on surface properties.

Influence of supramolecular self-assembly of oppositely charged Co- and Sn-porphyrins on the their spectral-luminescent properties in aqueous and aqueous micellar media of ionic surfactants

The processes of supramolecular self-assembly of Co(III)-meso-tetra-(4-trimethylammoniophenyl)porphyrin (CoP) and bis-(4-(1-imidazolyl)-phenol)-Sn(IV)-meso-tetra(4-sulfophenyl)porphyrin (SnP) were studied using electronic, steady-state and time-resolved fluorescence, one- and two-dimensional NMR spectroscopy in aqueous media. The main product of such interaction at a 2:1 M ratio of Co and Sn porphyrinates was found to be supramolecular porphyrin trimer CoP–SnP–CoP formed due to donor-acceptor, hydrogen and electrostatic interactions. The distinctive features of the resulting trimer are: significant distortion of the spatial structure of the Co-porphyrin fragments and strong quenching of the Sn-porphyrin macrocycles fluorescence (by 70 %). The distortion of the spatial structure of Co porphyrinates is in good agreement with the quantum chemical calculations as well as with the electronic and NMR spectroscopy data. Adding surfactants – both anionic (Cetyltrimethylammonium bromide - CTAB) and cationic (Sodium dodecyl sulfate - SDS) – even at concentrations below the critical micelle concentration (CMC) leads to the supramolecular trimer destruction. In solutions of ionic surfactants, supramolecular trimer destruction is caused by the interaction of either the peripheral macrocycles of the trimer (CoP) with SDS micelles (premicellar aggregates) or the central macrocycle (SnP) with CTAB micelles (premicellar aggregates). This process is prolonged and the rate of trimer destruction depends on the surfactant concentration. The sizes of the CTAB and SDS micelles solubilized with porphyrin molecules were established by dynamic light scattering. In general, surfactant additions lead to an increase in the SnP fluorescence at least due to the destruction of the supramolecular self-assembly, as well as due to the changes in the state of its microenvironment.

TADF Blue Emitters with Balanced π-Conjugation─Design, Synthesis, Spectral Characterization, and a Model OLED with 8-(5-(tert-Butyl)-1,3,4-oxadiazol-2-yl)-N,N-bis(4-(tert-butyl)phenyl)dibenzo[b,d]furan-2-amine.

Blue organic light-emitting diodes (OLED) suffer from relatively short lifetimes and a comparatively low lighting efficiency. One of the approaches to improving their characteristics is the development of luminophores with the potential for thermally activated delayed fluorescence (TADF). Herein, a set of donor-spacer-acceptor compounds with potential for TADF are designed, synthesized, and computationally and spectroscopically characterized. The excited state dynamics of the most prospective dye is monitored by time-resolved fluorescence and transient absorption spectroscopy. The experimental data are obtained and processed by a newly developed method and supplemented by quantum chemical calculations. The comprehensive approach allowed rationalization of the complex cascade-type photophysical behavior. The most promising emitter is included in an OLED displaying a blue color with a maximum EQE of 4.9% and negligible efficiency roll-off at higher luminance.

Modulation of Triton X-100 Aqueous Micelle Interface by Ionic Liquid: A Molecular Level Interaction Studied by Time-resolved Fluorescence Spectroscopy

Background: Self-assembly structure is an important area of research for understanding biological systems, owing to its resemblance to the membrane structure of the phospholipid bilayer. In a self-assembly medium, chemical reactions and chemical or physical processes are dramatically different than the bulk phase. Understanding this process in synthesizing self-assembly structures may allow us to explore various biological processes occurring in cell membranes. Objective: The study aimed to understand water dynamics in the TX-100 micellar interface via steady state and a time-resolved fluorescence spectroscopy study. The objective was also to determine the two different ionic liquids (ILs), namely 1-butyl-3-methyl imidazolium tetrafluoroborate ([bmim][BF4]) and 1-decyl-3-methyl imidazolium tetrafluoroborate ([dmim][BF4]), inducing surfactant aggregation changes at the molecular level. Also, the focus was on determining the hydration and its dynamics at the palisade layer of TX-100 micelle in the presence of two different ionic liquids. Methods: Steady state and time-resolved fluorescence spectroscopy have been used to study TX-100 micellar systems. Employing time-resolved spectroscopy, two chemical dynamic processes, solvation dynamics and rotational relaxation dynamics, have been studied to investigate structural changes in TX100 by adding ILs. Solvation dynamics was studied by measuring the time-dependent Stokes shift of the fluorescent probe. From the Stokes shift, time-resolved emission spectra were constructed to quantify the solvation dynamics. Also, using the polarization properties of light, time-resolved anisotropy was constructed to explore the rotation relaxation of the probe molecule. Results: The absorption and emission spectra of C-153 in TX-100 were red-shifted in the presence of both the ILs. Also, the C-153 experienced faster solvation dynamics and rotational relaxation with the addition of both ILs. In our previous study, we observed a significantly increased rate of solvation dynamics with the addition of [bmim][BF4] (J. Phys. Chem. B, 115, 6957-6963) [38]. However, with the addition of the same amount of [dmim][BF4], the IL rate of solvation enhancement was more pronounced than with [bmim][BF4]. The faster solvation and rotational relaxation have been found to be associated with the penetration of more free water at the TX100 micellar stern layer, leading to increased fluidity of the micellar interface. Conclusion: Upon incorporating ILs in TX100 micelle, substantially faster solvation dynamics of water as well as rotational relaxation dynamics of C-153 have been observed. By decreasing surfactant aggregations, [bmim][BF4] ILs facilitated more water molecules approaching the TX-100 micellar phase. On the other hand, [dmim][BF4] ILs comprising mixed micelles induced even more free water molecules at the palisade layer, yielding faster solvation dynamics in comparison to pure TX-100 micelle or TX100 micelle + [bmim][BF4] ILs systems. Time-resolved anisotropy study has also supported the finding and strengthened the solvation dynamics observation.

Characterization and classification of pathogenic bacteria using native fluorescence and spectral deconvolution.

Identification and classification of pathogenic bacterial strains is of current interest for the early treatment of diseases. In this work, protein fluorescence from eight different pathogenic bacterial strains were characterized using steady state and time resolved fluorescence spectroscopy. The spectral deconvolution method was also employed to decompose the emission contribution from different intrinsic fluorophores and extracted various key parameters, such as intensity, emission maxima, emission line width of the fluorophores, and optical redox ratio. The change in average lifetime values across different bacterial strains exhibits good statistical significance (p ≤ 0.01). The variations in the photophysical characteristics of bacterial strains are due to the different conformational states of the proteins. The stepwise multiple linear discriminate analysis of fluorescence emission spectra at 280 nm excitation across eight different bacterial strains classifies the original groups and cross validated group with 100% and 99.5% accuracy, respectively.

Trivalent lanthanide sorption onto illite in the presence of carbonate: A study combining thermodynamic sorption modeling, time-resolved laser fluorescence spectroscopy, and parallel factor analysis

Formation of aqueous actinide complexes with dissolved inorganic carbon (DIC) in groundwater decreases the sorption of actinides onto clay minerals in a geological disposal environment. Hence, the effect of DIC on actinide sorption must be evaluated quantitatively. In this study, batch sorption experiments using Eu(III) and Sm(III) as the chemical analogs of trivalent actinides were performed to assess their sorption behavior onto illite in the presence of DIC. In the studied pH region between 8.4 and 11, the sorption was significantly lower in the presence of DIC, with the largest effect being observed at pH 9–10. The effect increased with increasing DIC from 10 to 50 mM. A thermodynamic sorption model (TSM) was used to evaluate the results of batch sorption experiments. The experimental results can only be predicted when ternary-surface complexes with carbonate ions are considered. Moreover, information on the chemical forms of sorbed Eu on illite was obtained via time-resolved laser fluorescence spectroscopy (TRLFS) measurements. Parallel factor analysis of TRLFS data indicated the presence of two chemical species of Eu. These chemical species exhibited pH dependence, which is consistent with that of the surface species predicted using the TSM. In the presence of DIC, the dominant chemical species showed a long fluorescence lifetime and was inferred to coordinate with carbonate ions, supporting the validity of the TSM.

Mechanism understanding of PIKfyve inhibitor YM201636 with human serum albumin: Insights from molecular modeling and multiple spectroscopic techniques.

YM201636 is the potent PIKfyve inhibitor that is being actively investigated for liver cancer efficacy. In this study, computer simulations and experiments were conducted to investigate the interaction mechanism between YM201636 and the transport protein HSA. Results indicated that YM201636 is stably bound between the subdomains IIA and IIIA of HSA, supported by site marker displacement experiments. YM201636 quenched the endogenous fluorescence of HSA by static quenching since a decrease in quenching constants was observed from 7.74 to 2.39 × 104 M-1. UV-vis and time-resolved fluorescence spectroscopy confirmed the YM201636-HSA complex formation and this binding followed a static mechanism. Thermodynamic parameters ΔG, ΔH, and ΔS obtained negative values suggesting the binding was a spontaneous process driven by Van der Waals interactions and hydrogen binding. Binding constants ranged between 5.71 and 0.33 × 104 M-1, which demonstrated a moderately strong affinity of YM201636 to HSA. CD, synchronous, and 3D fluorescence spectroscopy revealed that YM201636 showed a slight change in secondary structure. The increase of Kapp and a decrease of PSH with YM201636 addition showed that YM201636 changed the surface hydrophobicity of HSA. The research provides reasonable models helping us further understand the transportation and distribution of YM201636 when it absorbs into the blood circulatory system.

Revisiting and Updating the Interaction between Human Serum Albumin and the Non-Steroidal Anti-Inflammatory Drugs Ketoprofen and Ketorolac.

Ketoprofen (KTF) and ketorolac (KTL) are among the most primarily used non-steroidal anti-inflammatory drugs (NSAIDs) in humans to alleviate moderate pain and to treat inflammation. Their binding affinity with albumin (the main globular protein responsible for the biodistribution of drugs in the bloodstream) was previously determined by spectroscopy without considering some conventional pitfalls. Thus, the present work updates the biophysical characterization of the interactions of HSA:KTF and HSA:KTL by 1H saturation-transfer difference nuclear magnetic resonance (1H STD-NMR), ultraviolet (UV) absorption, circular dichroism (CD), steady-state, and time-resolved fluorescence spectroscopies combined with in silico calculations. The binding of HSA:NSAIDs is spontaneous, endothermic, and entropically driven, leading to a conformational rearrangement of HSA with a slight decrease in the α-helix content (7.1% to 7.6%). The predominance of the static quenching mechanism (ground-state association) was identified. Thus, both Stern-Volmer quenching constant (KSV) and binding constant (Kb) values enabled the determination of the binding affinity. In this sense, the KSV and Kb values were found in the order of 104 M-1 at human body temperature, indicating moderate binding affinity with differences in the range of 0.7- and 3.4-fold between KTF and KTL, which agree with the previously reported experimental pharmacokinetic profile. According to 1H STD-NMR data combined with in silico calculations, the aromatic groups in relation to the aliphatic moiety of the drugs interact preferentially with HSA into subdomain IIIA (site II) and are stabilized by interactions via hydrogen bonding and hydrophobic forces. In general, the data obtained in this study have been revised and updated in comparison to those previously reported by other authors who did not account for inner filter corrections, spectral backgrounds, or the identification of the primary mathematical approach for determining the binding affinity of HSA:KTF and HSA:KTL.

Increasing Signal Intensity of Fluorescent Oligo-Labeled Antibodies to Enable Combination Multiplexing.

Full-spectrum flow cytometry has increased antibody-based multiplexing, yet further increases remain potentially impactful. We recently proposed how fluorescence multiplexing using spectral imaging and combinatorics (MuSIC) could do so using tandem dyes and an oligo-based antibody labeling method. In this work, we found that such labeled antibodies had significantly lower signal intensities than conventionally labeled antibodies in human cell experiments. To improve signal intensity, we tested moving the fluorophores from the original external (ext.) 5' or 3' end-labeled orientation to internal (int.) fluorophore modifications. Cell-free spectrophotometer measurements showed a ∼6-fold signal intensity increase of the new int. configuration compared to the previous ext. configuration. Time-resolved fluorescence and fluorescence correlation spectroscopy showed that the ∼3-fold brightness difference is due to static quenching most likely by the oligo or solution in the ext. configuration. Spectral flow cytometry experiments using peripheral blood mononuclear cells show int. MuSIC probe-labeled antibodies (i) retained increased signal intensity while having no significant difference in the estimated % of CD8+ lymphocytes and (ii) labeled with Atto488, Atto647, and Atto488/647 combinations can be demultiplexed in triple-stained samples. The antibody labeling approach is general and can be broadly applied to many biological and diagnostic applications where spectral detection is available.

Exploring the binding mechanism and esterase-like activity of human serum albumin with levofloxacin and its choline based conjugates: A biophysical approach

Human serum albumin (HSA) effectively binds to compounds having different molecular weight and thus facilitates their distribution in the living organisms. Thus, the binding interactions between a potential antibacterial drug (levofloxacin) and synthesized choline based levofloxacinate conjugates with HSA have been explored. The binding efficacy and mechanism were explored by utilizing different spectroscopic techniques; UV–Visible, steady state fluorescence, time resolved fluorescence and esterase-like activity. The interactions between the ligands and protein were electrostatic as well as hydrophobic in nature. The influence of different ligands having different alkyl chain shows quenching of the fluorescence emission of HSA. The spontaneous binding/quenching of HSA with ligands was static in nature, validated by steady state and time resolved fluorescence spectroscopy. Also, the impact of these ligands on the conformation of the native HSA structure was evaluated by using circular dichroism spectroscopy. In combination to the structural change study, the native protein functionality was observed (in terms of ‘esterase-like activity’) which has been found to be on lower side due to ligand binding. Further, we have performed the reverse study to check the impact of HSA on the fluorescent fluoroquinolone drug. The current study may prove helpful in elucidating the chemico-biological interactions which may prove useful in the pharmaceuticals, pharmacology, and different biochemistry fields.

2,6-Bis(5-(tert-butyl)-1H-pyrazol-3-yl)pyridine: Effects of the Peripheral Aliphatic Side Chain on the Coordination of Actinides(III) and Lanthanides(III).

To improve our understanding of the interaction mechanism in trivalent lanthanide and actinide complexes, studies with structurally different hard and soft donor ligands are of great interest. For that reason, the coordination chemistry of An(III) and Ln(III) with 2,6-bis(5-(tert-butyl)-1H-pyrazol-3-yl)pyridine (C4-BPP) has been explored. Time-resolved laser fluorescence spectroscopy (TRLFS) studies have revealed the formation of [Cm(C4-BPP)n]3+ (n = 1-3) (log β1' = 7.2 ± 0.4, log β2' = 10.1 ± 0.5, and log β3' = 11.8 ± 0.6) and [Eu(C4-BPP)m]3+ (m = 1-2) (log β1' = 4.9 ± 0.2 and log β2' = 8.0 ± 0.4). The absence of the [Eu(C4-BPP)3]3+ complex shows a more favorable complexation of Cm(III) over that of Eu(III). Additionally, complementary NMR measurements have been conducted to examine the M(III)-N bond in Ln(III) and Am(III) C4-BPP complexes. 15N NMR data have revealed notable differences in the chemical shifts of the coordinating nitrogen atoms between the Am(III) and Ln(III) complexes. In the Am(III) complex, the coordinating nitrogen atoms have shown a shift by 260 ppm, indicating a higher fraction of covalent bonding in the Am(III)-N bond compared with the Ln(III)-N bond. This observation aligns excellently with the differences in the stability constants obtained from TRLFS studies.

Increasing Signal Intensity of Fluorescent Oligo-Labeled Antibodies to Enable Combination Multiplexing.

Full-spectrum flow cytometry has increased antibody-based multiplexing, yet further increases remain potentially impactful. We recently proposed how fluorescence Multiplexing using Spectral Imaging and Combinatorics (MuSIC) could do so using tandem dyes and an oligo-based antibody labeling method. In this work, we found that such labeled antibodies had significantly lower signal intensity than conventionally-labeled antibodies in human cell experiments. To improve signal intensity, we tested moving the fluorophores from the original external (ext.) 5' or 3' end-labeled orientation to internal (int.) fluorophore modifications. Cell-free spectrophotometer measurements showed a ~6-fold signal intensity increase of the new int. configuration compared to the previous ext. configuration. Time-resolved fluorescence and fluorescence correlation spectroscopy showed that ~3-fold brightness difference is due to static quenching most likely by the oligo or solution in the ext. configuration. Spectral flow cytometry experiments using peripheral blood mononuclear cells show int. MuSIC probe-labeled antibodies (i) retained increased signal intensity while having no significant difference in the estimated % of CD8+ lymphocytes and (ii) labeled with Atto488, Atto647, and Atto488/647 combinations can be demultiplexed in triple-stained samples. The antibody labeling approach is general and can be broadly applied to many biological and diagnostic applications where spectral detection is available.

Ultrafast Energy Transfer in a Diatom Photosystem II Supercomplex.

The light-harvesting complexes (LHCs) of diatoms, specifically fucoxanthin-Chl a/c binding proteins (FCPs), exhibit structural and functional diversity, as highlighted by recent structural studies of photosystem II-FCP (PSII-FCPII) supercomplexes from different diatom species. The excitation dynamics of PSII-FCPII supercomplexes isolated from the diatom Thalassiosira pseudonana was explored using time-resolved fluorescence spectroscopy and two-dimensional electronic spectroscopy at room temperature and 77 K. Energy transfer between FCPII and PSII occurred remarkably fast (<5 ps), emphasizing the efficiency of FCPII as a light-harvesting antenna. The presence of long-wavelength chlorophylls may further help concentrate excitations in the core complex and increase the efficiency of light harvesting. Structure-based calculations reveal remarkably strong excitonic couplings between chlorophylls in the FCP antenna and between FCP and the PSII core antenna that are the basis for the rapid energy transfer.

Fiber optic-based integrated system for in vivo multiscale pharmacokinetic monitoring.

This paper presents the development of a fiber-optic-based fluorescence detection system for multi-scale monitoring of drug distribution in living animals. The integrated system utilized dual laser sources at the wavelengths of 488 nm and 650 nm and three photomultiplier channels for multi-color fluorescence detection. The emission spectra of fluorescent substances were tracked using the time-resolved fluorescence spectroscopy module to continuously monitor their blood kinetics. The fiber bundle, consisting of 30,000 optic filaments, was designed for wide-field mesoscopic imaging of the drug's interactions within organs. The inclusion of a gradient refractive index (GRIN) lens within the setup enabled fluorescence confocal laser scanning microscopy to visualize the drug distribution at the cellular level. The system performance was verified by imaging hepatic and renal tissues in mice using cadmium telluride quantum dots (CdTe QDs) and R3. By acquiring multi-level images and real-time data, our integrated system underscores its potential as a potent tool for drug assessment, specifically within the realms of pharmacokinetic and pharmacodynamic investigations.

References for Small Fluorescence Quantum Yields.

Three compounds with fluorescence quantum yields in the range of 10- 5 to 10- 4 and emission spectra covering the UV/Vis spectral range are suggested as new references for the determination of small fluorescence quantum yields. The compounds are thymidine (dT) in water, dibenzoylmethane (DBM) in ethanol, and malachite green chloride (MG) in water, representing the blue, green, and red regions of the spectrum, respectively. All compounds are easily handled, photostable, and commercially available. Furthermore, these compounds exhibit a mirror-image symmetry between their absorption and fluorescence spectra. This symmetry, along with closely aligned fluorescence excitation and absorption spectra, confirms that the observed emissions originate from the compounds themselves. The fluorescence quantum yields were determined via a relative approach as well as Strickler-Berg analysis in conjunction with time resolved fluorescence spectroscopy. Within the respective error margins, the two approaches yielded identical results.