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Organic small‐molecule fluorophores, characterized by flexible chemical structure and adjustable optical performance, have shown tremendous potential in biosensing. However, classical organic fluorophore motifs feature large overlap between excitation and emission spectra, leading to the requirement of advanced optical set up to filter desired signal, which limits their application in scenarios with simple settings. Here, a series of wavelength‐tunable small‐molecule fluorescent dyes (PTs) bearing simple organic moieties have been developed, which exhibit Stokes shift up to 262 nm, molar extinction coefficients ranged 30,000–100,000 M−1 cm−1, with quantum yields up to 54.8 %. Furthermore, these dyes were formulated into fluorescent nanoparticles (PT‐NPs), and applied in lateral flow assay (LFA). Consequently, limit of detection for SARS‐CoV‐2 nucleocapsid protein reached 20 fM with naked eye, a 100‐fold improvement in sensitivity compared to the pM detection level for colloidal gold‐based LFA. Besides, combined with loop‐mediated isothermal amplification (LAMP), the LFA system achieved the visualization of single copy level nucleic acid detection for monkeypox (Mpox). [ABSTRACT FROM AUTHOR]

We report synthesis of a library of diamino‐terephthalonitrile based single‐benzene fluorophores (SBFs). These SBFs contain a reactive terminal linker group. They open up the possibility of marrying their excellent fluorescent property with various biogenic units, leading to value‐added SBFs. The solvatofluorochromism behavior of these newly synthesized SBFs was also studied. The emission enhances and blue‐shifts in case of non‐polar solvents and vice versa in the polar solvents, implying the environmental sensitivity of these fluorophores. Photostability test was also done to test for the stability of the emission of these fluorophores with time and the fluorophores were found to be stable. We also observed time‐dependent fluorescence enhancement of one of our SBFs in presence of reactive biogenic amines. Overall, the generation of these novel molecules opens up new opportunities towards bioconjugated fluorophores. [ABSTRACT FROM AUTHOR]

Second near‐infrared (NIR‐II) bioimaging technology shows broad application prospects in the field of biomedical due to its significant superiority in deep tissue penetration and high signal‐to‐background ratio. Among them, organic small molecule fluorophores have received progressively more consideration due to their low biotoxicity, high biocompatibility, rapid clearance, and good modification properties. As the key acceptor core, 2, 1, 3‐benzothiadiazole (BTD) derivative plays an essential role in the construction of donor‐acceptor‐donor type NIR‐II fluorophores. This mini‐review summarizes the research development of NIR‐II small molecule fluorophores based on BTD derivatives in the field of bioimaging, focusing on molecular design and photophysical properties, and introduces a part of the applications in the biological field. Furthermore, the challenges of BTD‐based NIR‐II fluorophores in the future development and application are discussed. [ABSTRACT FROM AUTHOR]

Bacterial infections have emerged as the leading causes of mortality and morbidity worldwide. Herein, we developed a dual‐channel fluorescence "turn‐on" sensor array, comprising six electrostatic complexes formed from one negatively charged poly(para‐aryleneethynylene) (PPE) and six positively charged aggregation‐induced emission (AIE) fluorophores. The 6‐element array enabled the simultaneous identification of 20 bacteria (OD600=0.005) within 30s (99.0 % accuracy), demonstrating significant advantages over the array constituted by the 7 separate elements that constitute the complexes. Meanwhile, the array realized different mixing ratios and quantitative detection of prevalent bacteria associated with urinary tract infection (UTI). It also excelled in distinguishing six simulated bacteria samples in artificial urine. Remarkably, the limit of detection for E. coli and E. faecalis was notably low, at 0.000295 and 0.000329 (OD600), respectively. Finally, optimized by diverse machine learning algorithms, the designed array achieved 96.7 % accuracy in differentiating UTI clinical samples from healthy individuals using a random forest model, demonstrating the great potential for medical diagnostic applications. [ABSTRACT FROM AUTHOR]

STED nanoscopy allows for the direct observation of dynamic processes in living cells and tissues with diffraction-unlimited resolution. Although fluorescent proteins can be used for STED imaging, these labels are often outperformed in photostability by organic fluorescent dyes. This feature is especially crucial for time-lapse imaging. Unlike fluorescent proteins, organic fluorophores cannot be genetically fused to a target protein but require different labeling strategies. To achieve simultaneous imaging of more than one protein in the interior of the cell with organic fluorophores, bioorthogonal labeling techniques and cell-permeable dyes are needed. In addition, the fluorophores should preferentially emit in the red spectral range to reduce the potential phototoxic effects that can be induced by the STED light, which further restricts the choice of suitable markers. In this work, we selected five different cell-permeable organic dyes that fulfill all of the above requirements and applied them for SPIEDAC click labeling inside living cells. By combining click-chemistry-based protein labeling with other orthogonal and highly specific labeling methods, we demonstrate two-color STED imaging of different target structures in living specimens using different dye pairs. The excellent photostability of the dyes enables STED imaging for up to 60 frames, allowing the observation of dynamic processes in living cells over extended time periods at super-resolution. [ABSTRACT FROM AUTHOR]

Currently, the use of synthetic pigments in foods is restricted since synthetic pigments are proven and suspected to be harmful to human health. Phycobiliproteins (PBPs), existed in phycobilisomes (PBSs) of algae, are a kind of pigment-proteins with intense color. The specific color of PBPs (red and blue) is given by the water-soluble open-chained tetrapyrrole chromophore (phycobilin) that covalently attaches to the apo-protein via thioether linkages to cysteine residues. According to the spectral characteristics of PBPs, they can be categorized as phycoerythrins (PEs), phycocyanins (PCs), allophycocyanins (APCs), and phycoerythrocyanins (PECs). PBPs can be used as natural food colorants, fluorescent substances, and bioactive ingredients in food applications owing to their color characteristics and physiological activities. This paper mainly summarizes the extraction and purification methods of the PBPs and reviews their characteristics and applications. Moreover, the use of several strategies such as additives, microencapsulation, electrospray, and cross-linking to improve the stability and bioavailability of PBPs as well as the future outlooks of PBPs as natural colorants in food commercialization are elucidated. [ABSTRACT FROM AUTHOR]

This article presents a study on the synthesis and characterization of three new donor-acceptor conjugates that have the ability to sense changes in pH and detect fluoride ions. The molecules were analyzed using various techniques and their optical properties were investigated. The compounds showed solvatochromic effects on their fluorescence spectra and demonstrated pH sensitivity, with a turn-on fluorescence response towards acidic pH. They also selectively bound to fluoride ions, resulting in a fluorescence response. The article provides detailed experimental procedures and characterization data for each compound. [Extracted from the article]

The development of near‐infrared (NIR) fluorophores that have both excellent chemical stability and photostability, as well as efficient cell permeability, is highly demanded. In this study, we present phospha‐rhodamine (POR) dyes which display significantly improved performance for protein labeling. This is achieved by incorporating a 2‐carboxy‐3‐benzothiophenyl group at the 9‐position of the xanthene scaffold. The resulting cis and trans isomers were successfully isolated and structurally characterized using X‐ray diffraction. The HaloTag ligand conjugates of the two isomers exhibited different staining abilities in live cells. While the cis isomer showed non‐specific accumulation on the organelle membranes, the trans isomer selectively labeled the HaloTag‐fused proteins, enabling the long‐term imaging of cell division and the 5‐color imaging of cell organelles. Molecular dynamics simulations of the HaloTag ligand conjugates within the lipid membrane suggested that the cis isomer is more prone to forming oligomers in the membrane. In contrast, the oligomerization of the trans isomer is effectively suppressed by its interaction with the lipid molecules. By taking advantage of the superior labeling performance of the trans isomer and its NIR‐emissive properties, multi‐color time‐lapse super‐resolution 3D imaging, namely super‐resolution 5D‐imaging, of the interconnected network between the endoplasmic reticulum and microtubules was achieved in living cells. [ABSTRACT FROM AUTHOR]

In this work, we developed a rapid and sensitive label-free ratiometric fluorescent (FL) probe for the detection of bleomycin (BLM). The probe consists of a DNA sequence (D6) and two fluorophore groups, 2-amino-5,6,7-trimethyl-1,8-naphthalene (ATMND) and SYBR Green I (SGI). The D6 sequence could be folded into a three-way junction structure containing a C–C mismatch position in the junction pocket. The unique "Y" structure not only could entrap ATMND in the mismatch pocket with high affinity, leading to FL quenching at 408 nm, but also embed SGI in the grooves of the double-stranded portion, resulting in FL enhancement at 530 nm. In the presence of BLM–Fe(II), the "Y" structure of D6 was destroyed due to the specific cleavage of the BLM recognition site, the 5′-GT-3′ site in D6. This caused the release of ATMND and SGI and thus the ratiometric signal change of FL enhancement by ATMND and FL quenching by SGI. Under optimal conditions, the ratiometric probe exhibited a linear correlation between the intensity ratio of F408/F530 and the concentration of BLM in the range of 0.5–1000 nM, with a detection limit of 0.2 nM. In addition, the probe was applied to detect BLM in human serum samples with satisfactory results, indicating its good clinical application potential. [ABSTRACT FROM AUTHOR]

Photoluminescence (PL) color can be tuned by mixing fluorophores emitting the three primary colors in an appropriate ratio. When color tuning is achieved on a single substrate, we can simplify device structures. We demonstrated that nanographenes (NGs), which are graphene fragments with a size of tens of nanometers, could be utilized as carriers of fluorophores. The addition of red‐ and blue‐light‐emitting fluorophores on the edge successfully reproduced the purple light. The relative PL intensities of the fluorophores could be regulated by the excitation wavelength, enabling multicolor emission between blue and red light. Owing to the triphenylamine units of the fluorophores, the NGs showed PL enhancement due to aggregation. This characteristic was valuable for the fabrication of solid polymer materials. Specifically, the functionalized NGs can be dispersed into polyvinylidene difluoride. The resultant polymer films emitted red, blue, and purple color. Our study demonstrated the potential applicability of NGs for fluorophore carriers capable of reproducing intermediate colors of light. [ABSTRACT FROM AUTHOR]

Fluorescence‐detected linear dichroism (FD‐LD) enables one to collect linear dichroism spectra for oriented fluorophores in the presence of other absorbing species and light scattering. The experiment proceeds by scanning the excitation wavelength and using a filter to collect only emitted photons from the fluorophore. Thus, it has the potential to give data with enhanced selectivity and quality. By using a synchrotron radiation light source and fluorescence‐detection, we show data for a range of fluorophores in different orienting environments. Film and flow‐oriented FD‐LD spectra were collected down to 170 nm. Even for flow‐oriented liposomes, we have data collected down to 210 nm. For strongly scattering samples, for example, liposomes, FD‐LD has the clear advantage that scattering is absent for the longer wavelength fluorescence photons. The collimated and smaller beam size of the synchrotron radiation also gives rise to sharper and more well‐defined features in the spectra. [ABSTRACT FROM AUTHOR]

The design and development of ultra‐accurate probe is of great significance to chemical sensing in complex practical scenarios. Here, a self‐accelerating naphthalimide‐based probe with fast response and high sensitivity toward hydrogen peroxide (H2O2) is designed. By coupling with the specially selected upconversion nanoparticles (UCNPs), an ultra‐accurate colorimetric‐fluorescent‐upconversion luminescence (UCL) tri‐mode platform is constructed. Owing to the promoted reaction process, this platform demonstrates rapid response (< 1 s), an ultra‐low detection limit (4.34 nM), and superb anti‐interferent ability even in presence of > 21 types of oxidants, explosives, metallic salts, daily compounds, colorful or fluorescent substances. In addition, the effectiveness of this design is further verified by a sponge‐based sensing chip loaded with the UCNPs/probe in recognizing trace H2O2 vapor from interferents with the three characteristic colors existing simultaneously. The proposed design of probe and tri‐mode visualization detection platform is expected to open up a brand‐new methodology for ultra‐accurate sensing. [ABSTRACT FROM AUTHOR]

A 49-year-old man attended an oral health campaign for a preliminary screening of potentially malignant lesions using a fluorescent developer substance. At the time of examination, he displayed no apparent symptoms. However, upon questioning, the man disclosed having uncontrolled diabetes and hypertension, as well as a history of alcohol consumption and tobacco smoking. He reported no family history of cancer, no prior surgeries or dental procedures, and no dental visits in the past decade. Clinical examination of the oral cavity revealed poor oral health, tooth loss, and mobility, with bleeding noted in some posterior teeth. Application of the fluorescent substance revealed areas with discoloration distinct from the oral mucosa, leading to a presumptive positive diagnosis of potentially malignant oral lesions. Additionally, dental tartar and floating dentobacterial plaque were identified. The substance's use also aids in the identification of microbial structures, facilitating easy detection. [ABSTRACT FROM AUTHOR]

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Creation of new fluorophores is important for understanding the structure–property relationship, by which the required optical properties are likely to be attained. Herein, through theory calculation, it is found that furan‐modified thiadiazolo quinoxaline acting as an electron acceptor can endow donor–acceptor–donor (D–A–D) type second near‐infrared (NIR‐II) fluorophores with longer emission wavelength than the other thiadiazolo quinoxaline‐based acceptors containing pyridine, pyrrole, thiophene, and phenyl groups, respectively. On the basis of this theoretical prediction, a D–A–D type NIR‐II fluorophore with 6,7‐di(furan‐2‐yl)‐[1,2,5]thiadiazolo[3,4‐g] quinoxaline (DFTQ) as the acceptor and dithieno[3,2‐b:2′,3′‐d]pyrrole (DTP) as the donor is designed and synthesized, and the aggregation‐induced emission (AIE) function is further achieved by introducing the AIE units of tetraphenylethylene (TPE) and triphenylamine (TPA), respectively, totally forming three NIR‐II fluorophores DFTQ–DTP, DFTQ–DTPE, and DFTQ–DTPA. For biological applications, the fluorophores are encapsulated by amphiphilic DSPE–PEG2000 to generate water‐dispersible nanoparticles (NPs). Almost the whole emission of each of the NPs falls into the NIR‐II spectral range, with part emission beyond 1300 nm. By using DFTQ–DTPA NPs as the contrast and photothermal therapy (PTT) agent, high‐resolution in vivo fluorescence imaging is achieved in the greater than 1300 nm window, and their good performance in photoacoustic imaging and high tumor PTT efficacy in tumor‐bearing mice are also demonstrated. Taken together, this work mainly provides a strong electron acceptor for constructing long‐emitting fluorophores, and by using the electron acceptor, a AIE fluorophore with desirable quantum yield (QY) and photothermal conversion efficienciy (PCE) is synthesized and demonstrated to be promising in fluorescence/photoacoustic imaging and PTT. [ABSTRACT FROM AUTHOR]

Raman scattering can provide information on molecular fingerprints, which have been widely applied in various fields of material science and nanobiotechnology. Notably, low interference with water molecules in obtaining the Raman spectra between 500 and 2000 cm−1 made it a powerful spectroscopic tool in biology, such as imaging and signaling for a living cell. To be a robust tool for cell biology, the performance of obtaining molecular-specific information with high sensitivity, high resolution in real time, and without inducing cell damage is strongly required. The conventional fluorescence-based method has been suffered from the rapid photobleaching of organic fluorophores and the lack of molecular information. In contrast, Raman scattering is a promising spectroscopic tool to acquire cellular information, and the extremely low signal intensity of Raman scattering could be amplified by incorporating the plasmonic nanomaterials. Along with the fundamental research focus on surface-enhanced Raman scattering (SERS), the practical approaches of SERS for cellular imaging as a new tool for drug screening and monitoring cellular signals have been extensively explored based on new optical setups and new designing strategies for the nanostructures. Diverse nanostructure and surface chemistry for targeting or sensing have been played pivotal roles in acquiring cellular information and high resolution cell imaging. In this regard, this review focused on the recent advances of SERS-based technologies for a live cell imaging investigated such as potential drug screening, signaling for chemicals or biomolecules in cell, in situ sensing, and high spatiotemporal resolution. [ABSTRACT FROM AUTHOR]

Caged compounds are frequently used in life science research. However, the light used to activate them is commonly absorbed and scattered by biological materials, limiting their use to basic research in cells or small animals. In contrast, hard X‐rays exhibit high bio‐permeability due to the difficulty of interacting with biological molecules. With the main goal of developing X‐ray activatable caged compounds, azo compounds are designed and synthesized with a positive charge and long π‐conjugated system to increase the reaction efficiency with hydrated electrons. The azo bonds in the designed compounds are selectively cleaved by X‐ray, and the fluorescent substance Diethyl Rhodamine is released. Based on the results of experiments and quantum chemical calculations, azo bond cleavage is assumed to occur via a two‐step process: a two‐electron reduction of the azo bond followed by N─N bond cleavage. Cellular experiments also demonstrate that the azo bonds can be cleaved intracellularly. Thus, caged compounds that can be activated by an azo bond cleavage reaction promoted by X‐ray are successfully generated. [ABSTRACT FROM AUTHOR]

Fluorescence spectroscopy proves to be a powerful and versatile tool in scientific research, demanding ongoing advancements of fluorescence probes to cater to a broad range of applications. The particular interest lies in the development of fluorophores with multiplexing capability, emitting from more than one excited state depending on the polarity and viscosity of the surrounding medium. Here, we present a variety of novel dual planarized intramolecular charge transfer (PLICT) emitters based on cyano‐1‐aminoindole structures. We have chosen a combination of experimental studies, theoretical calculations and the creation of a unique fluorophore cataloging map. This map serves as a useful tool for empirically determining the origin of both fluorescence bands. [ABSTRACT FROM AUTHOR]

A catalytic carbonylative method for the direct, multicomponent synthesis of indolizine fluorophores has been developed. The process is based on the PdI2/KI‐catalyzed oxidative aminocarbonylation of 2‐(pyridin‐2‐yl)pent‐4‐yn‐1‐carbonyl compounds leading to the corresponding 2‐ynamide intermediates, followed in situ by 5‐exo‐dig cyclization (by amine‐promoted conjugate addition of the pyridine ring nitrogen) and aromatization, to give the finally isolated N,N‐disubstituted 2‐(indolizin‐3‐yl)acetamides. Reactions are carried out under relatively mild conditions (100 °C under 20 atm of a 4 : 1 mixture CO−air for 6–15 h) in MeCN as the solvent and with a low catalyst loading (0.33 mol% PdI2), in the presence of 0.5 equiv. of KI and 3 equiv. of a secondary amine. The optical properties of representative 2‐(indolizin‐3‐yl)acetamide products have also been investigated. [ABSTRACT FROM AUTHOR]

The pulmonary vascular system plays a crucial role in maintaining normal physiological functions, and perturbations in this network often serve as indicators for various fatal diseases. Thus, accurate mapping and assessment of the intricate anatomical details of pulmonary vasculature is essential for the investigation of the underlying mechanism of these diseases. Yet it is considered a tough challenge as traditional imaging techniques offer limited representations of the vasculature network in the lung, while optical imaging methods face limitations from tissue depth. To overcome these obstacles, in this study, an AIENP‐reinforced DISCO method, for whole‐tissue 3D reconstruction of pulmonary capillaries is presented. Combining AIENPs, hydrogel‐enhanced scaffolds, and solvent‐based DISCO procedures, the method successfully visualizes the entire network of mouse pulmonary capillaries with a significantly shortened timeframe and cost. The whole process including labeling and clearing takes 6 days and it costs ≈ 5 USD to stain the lung vasculature of an adult mouse. Moreover, the study provides valuable insights for detecting pulmonary vascular abnormalities. This fast and cost‐effective technique opens new avenues for developing better fluorophores compatible with tissue optical clearing and offers insights for in‐depth research on pulmonary pathophysiology. [ABSTRACT FROM AUTHOR]

Au(III) bis(pyrrolide-imine) chelates are emerging as a class of versatile, efficacious metallodrug candidates. Here, we synthesised two enantiopure chiral ligands H2L1 and H2L2 (tetradentate cyclohexane-1,2-diamine-bridged bis(pyrrole-imine) derivatives). Metallation of the ligands with Au(III) afforded the chiral cationic complexes AuL1 and AuL2. The in vitro cytotoxicities of AuL1 and AuL2 determined in the NCI-60 single-dose drug screen were 56.5% and 89.1%, respectively. AuL1 was subsequently selected for a five-dose NCI-60 screen, attaining GI50, IC50, and LC50 values of 4.7, 9.3 and 39.8 μM, respectively. Hierarchical cluster analysis of the NCI-60 data indicated that the profile for AuL1 was similar to that of vinblastine sulfate, a microtubule-targeting vinca alkaloid. Reactions of AuL1 with glutathione (GSH) in vitro confirmed its susceptibility to reduction, Au(III) → Au(I), by intracellular thiols. Because human serum albumin (HSA) is responsible for transporting clinically deployed and investigational drugs, we studied the uptake of AuL1 and AuL2 by HSA to delineate how chirality impacts their protein-binding affinity. Steady-state fluorescence quenching data acquired on the native protein and data from site-specific probes showed that the compounds bind at sites close enough to Trp-214 (subdomain IIA) of HSA to quench the fluorophore. The bimolecular quenching rate constants, Kq, were ca. 102 times higher than the maximum diffusion-controlled collision constant of a biomolecule in water (1010 M−1 s−1), confirming that static fluorescence quenching was the dominant mechanism. The Stern–Volmer constants, KSV, were ∼104 M−1 at 37 °C, while the affinity constants, Ka (37 °C), measured ∼2.1 × 104 M−1 (AuL1) and ∼1.2 × 104 M−1 (AuL2) for enthalpy-driven ligand uptake targeting Sudlow's site I. Although far- and near-UV CD spectroscopy indicated that both complexes minimally perturb the secondary and tertiary structure of HSA, substantial shifts in the CD spectra were recorded for both protein-bound ligands. This study highlights the role of chirality in determining the cytotoxicity profiles and protein binding behaviour of enantiomeric Au(III) chelates. [ABSTRACT FROM AUTHOR]

Förster resonance energy transfer (FRET) probes are a promising tool for studying numerous biochemical processes. In this paper, we show the application of the FRET phenomenon to observe the micelle formation from surfactants, micelles self-assembling from chitosan grafted with fatty acid (oleic—OA, or lipoic—LA), cross-linking of SH groups in the micelle's core, and inclusion and release of the model drug cargo from the micelles. Using the carbodiimide approach, amphiphilic chitosan-based polymers with (1) SH groups, (2) crosslinked with S-S between polymer chains, and (3) without SH and S-S groups were synthesized, followed by characterization by FTIR and NMR spectroscopy. Two pairs of fluorophores were investigated: 4-methylumbelliferon-trimethylammoniocinnamate—rhodamine (MUTMAC–R6G) and fluorescein isothiocyanate—rhodamine (FITC–R6G). While FITC–R6G has been described before as an FRET-producing pair, for MUTMAC–R6G, this has not been described. R6G, in addition to being an acceptor fluorophore, also serves as a model cytostatic drug in drug-release experiments. As one could expect, in aqueous solution, FRET effect was poor, but when exposed to the micelles, both MUTMAC–R6G and FITC–R6G yielded a pronounced FRET effect. Most likely, the formation of micelles is accompanied by the forced convergence of fluorophores in the hydrophobic micelle core by a donor-to-acceptor distance (r) significantly closer than in the aqueous buffer solution, which was reflected in the increase in the FRET efficiency (E). Therefore, r(E) could be used as analytical signal of the micelle formation, including critical micelle concentration (CMC) and critical pre-micelle concentration (CPMC), yielding values in good agreement with the literature for similar systems. We found that the r-function provides analytically valuable information about the nature and mechanism of micelle formation. S-S crosslinking between polymer chains makes the micelle more compact and stable in the normal physiological conditions, but loosens in the glutathione-rich tumor microenvironment, which is considered as an efficient approach in targeted drug delivery. Indeed, we found that R6G, as a model cytostatic agent, is released from micelles with initial rate of 5%/h in a normal tissue microenvironment, but in a tumor microenvironment model (10 mM glutathione), the release of R6G from S-S stitched polymeric micelles increased up to 24%/h. Drug-loading capacity differed substantially: from 75–80% for nonstitched polymeric micelles to ~90% for S-S stitched micelles. Therefore, appropriate FRET probes can provide comprehensive information about the micellar system, thus helping to fine-tune the drug delivery system. [ABSTRACT FROM AUTHOR]

Organic fluorophores are indispensable tools in cells, tissue and in vivo imaging, and have enabled much progress in the wide range of biological and biomedical fields. However, many available dyes suffer from insufficient performances, such as short absorption and emission wavelength, low brightness, poor stability, small Stokes shift, and unsuitable permeability, restricting their application in advanced imaging technology and complex imaging. Over the past two decades, many efforts have been made to improve these performances of fluorophores. Starting with the luminescence principle of fluorophores, this review clarifies the mechanisms of the insufficient performance for traditional fluorophores to a certain extent, systematically summarizes the modified approaches of optimizing properties, highlights the typical applications of the improved fluorophores in imaging and sensing, and indicates existing problems and challenges in this area. This progress not only proves the significance of improving fluorophores properties, but also provide a theoretical guidance for the development of high‐performance fluorophores. [ABSTRACT FROM AUTHOR]

Life science has progressed with applications of fluorescent probes—fluorophores linked to functional units responding to biological events. To meet the varied demands across experiments, simple organic reactions to connect fluorophores and functional units have been developed, enabling the on‐demand selection of fluorophore‐functional unit combinations. However, organic synthesis requires professional equipment and skills, standing as a daunting task for life scientists. In this study, we present a simple, fast, and convenient strategy for probe preparation: co‐aggregation of hydrophobic molecules. We focused on tetrazine—a difficult‐to‐prepare yet useful functional unit that provides effective bioorthogonal reactivity and strong fluorogenicity. Simply mixing the tetrazine molecules and aggregation‐induced emission (AIE) luminogens in water, co‐aggregation is induced, and the emission of AIE luminogens is quenched. Subsequent click reaction bioorthogonally turns on the emission, identifying these coaggregates as fluorogenic probes. Thanks to this bioorthogonal fluorogenicity, we established a new time‐gated fluorescence bioimaging technique to distinguish overlapping emission signals, enabling multi‐organelle imaging with two same‐color fluorophores. Our study showcases the potential of this co‐aggregation method for the on‐demand preparation of fluorescent probes as well as protocols and molecular design principles in this approach, offering an effective solution to evolving needs in life science research. [ABSTRACT FROM AUTHOR]

We introduce MINFLUX localization with interferometric illumination through opposing objective lenses for maximizing the attainable precision in 3D-localization of single inelastic scatterers, such as fluorophores. Our 4Pi optical configuration employs three sequentially tilted counter-propagating beam pairs for illumination, each providing a narrow interference minimum of illumination intensity at the focal point. The localization precision is additionally improved by adding the inelastically scattered or fluorescence photons collected through both objective lenses. Our 4Pi configuration yields the currently highest precision per detected photon among all localization schemes. Tracking gold nanoparticles as non-blinking inelastic scatterers rendered a position uncertainty <0.4 nm³ in volume at a localization frequency of 2.9 kHz. We harnessed the record spatio-temporal precision of our 4Pi MINFLUX approach to examine the diffusion of single fluorophores and fluorescent nanobeads in solutions of sucrose in water, revealing local heterogeneities at the nanoscale. Our results show the applicability of 4Pi MINFLUX to study molecular nano-environments of diffusion and its potential for quantifying rapid movements of molecules in cells and other material composites. [ABSTRACT FROM AUTHOR]

The recent discovery of blue fluorophores with high quantum yields based on pyridone structures inspired the development of new low‐molecular‐weight fluorophores with bright emissions at tunable wavelengths, which are highly attractive for various applications. In this study, we propose a rational design strategy for 2‐pyridone‐based fluorophores with bright emissions at long wavelengths. With a detailed understanding of the positional substitution effects on each carbon atom of the 2‐pyridone core, we developed a bright blue fluorophore (λabs=377 nm; λem=433 nm; ϵ=13,200 M−1 cm−1; ϕF=88 %) through C3‐aryl and C4‐ester substitutions followed by cyclization. Furthermore, by applying the intramolecular charge transfer (ICT) principle, we invented a bright green fluorophore through C3‐ and C4‐diester and C6‐aryl substitutions. The ICT fluorophore based on the pyridone structure shows large molar absorptivity (ϵ=20,100 M−1 cm−1), longer emission wavelength (λem=539 nm), high emission quantum yield (ϕF=74 %), and large Stokes shift (Δv=5720 cm−1), which are comparable to those of practical fluorescent probes. [ABSTRACT FROM AUTHOR]

Near‐infrared (NIR) fluorophores are highly desirable for super‐resolution fluorescence microscopy; however, their optical properties are typically relatively poor, with low photo‐intensity (mean photon number below 1000), and unfavorably high on‐off blinking duty cycle. In this work, the synthesis and characterization of tetrabenzodinaphthopyranthren (TBDNP) derivative as a novel nanographene that exhibits NIR emission in the range of 800–1000 nm with a photoluminescence quantum yield of 14% are reported. TBDNP demonstrated excellent self‐blinking properties, in particular, mean photon number of >3000 and a low on–off‐duty cycle (≈10−4), which are superior to reported NIR dyes, and can be successfully employed for the NIR super‐resolution imaging. Moreover, TBDNP serves also as a chiral fluorophore, showing notable chiroptical activities from 300 to 850 nm with a high dissymmetry factor (gabs) of 0.032 at 574 nm. [ABSTRACT FROM AUTHOR]

Real-time monitoring of biocatalytic-based processes is significantly improved and simplified when they can be visualized. Visual monitoring can be achieved by integrating a fluorescent unit with the biocatalyst. Herein, we outline the design strategies of fluorescent probes for monitoring biocatalysis: (1) probes for monitoring biocatalytic transfer: γ-glutamine is linked to the fluorophore as both a recognition group and for intramolecular charge transfer (ICT) inhibition; the probe is initially in an off state and is activated via the transfer of the γ-glutamine group and the release of the free amino group, which results in restoration of the "Donor–π–Acceptor" (D–π–A) system and fluorescence recovery. (2) Probes for monitoring biocatalytic oxidation: a propylamine is connected to the fluorophore as a recognition group, which cages the hydroxyl group, leading to the inhibition of ICT; propylamine is oxidized and subsequently β-elimination occurs, resulting in exposure of the hydroxyl group and fluorescence recovery. (3) Probes for monitoring biocatalytic reduction: a nitro group attached to a fluorophore as a fluorescence quenching group, this is converted to an amino group by catalytic reduction, resulting in fluorescence recovery. (4) Probes for monitoring biocatalytic hydrolysis: β- D -galactopyranoside or phosphate acts as a recognition group attached to hydroxyl groups of the fluorophore; the subsequent biocatalytic hydrolysis reaction releases the hydroxyl group resulting in fluorescence recovery. Following these 4 mechanisms, fluorophores including cyanine, coumarin, rhodamine, and Nile-red, have been used to develop systems for monitoring biocatalytic reactions. We anticipate that these strategies will result in systems able to rapidly diagnose and facilitate the treatment of serious diseases. [ABSTRACT FROM AUTHOR]

In this work, a novel organic receptor, CPI [(E)-3-(4-(9H-carbazol-9-yl)phenyl)-2-(1H-benzo[d]imidazol-2-yl)acrylonitrile], was rationally designed and successfully fabricated for selective and sole recognition of CN− ions over other competitive anions through an obvious chromogenic and ratiometric emission change in DMSO. The distinct and prominent color change upon the addition of CN− can be attributed to the typical ICT process, which is induced by the deprotonation of acidic NH protons in the imidazole moiety. The sensor displayed strong solvatochromic effects in commonly used organic solvents such as n-hexane, toluene, diethyl ether, DCM, THF, DMF and DMSO. The chemical structure of the sensor was characterized by single-crystal X-ray diffraction, 1HNMR, 13CNMR, IR and mass spectroscopy. Significantly, the probe can function as a fluorescence-based sensor for the efficient detection of low-level water in organic solvents. The solid-state emission properties of CPI were successfully applied to recognise cyanide in a solid-state platform with naked eye-visualized distinct color change. The probe can be made reusable by adding TFA into the CN− treated probe solution. The detection limit of CPI towards CN− was determined to be 4.48 × 10−8 M. More importantly, the sensor is capable of detecting CN− in food samples and has been employed for wastewater treatment. Besides, easy-to-prepare CPI-coated test strips provide a simple, reusable and easy-to-handle protocol for the qualitative identification of CN− conveniently. Finally, density functional theory and time-dependent density functional theory were performed to verify the experimental outcomes theoretically. [ABSTRACT FROM AUTHOR]

The autofluorescence-spectral imaging (ASI) technique is based on the lightemitting ability of natural fluorophores. Soybean genotypes showing contrasting tolerance to pre-germination anaerobic stress can be characterized using the photon absorption and fluorescence emission of natural fluorophores occurring in seed coats. In this study, tolerant seeds were efficiently distinguished from susceptible genotypes at 405 nm and 638 nm excitation wavelengths. ASI approach can be employed as a new marker for the detection of photonemitting compounds in the tolerant and susceptible soybean seed coats. Furthermore, the accuracy of rapid characterization of genotypes using this technique can provide novel insights into soybean breeding. [ABSTRACT FROM AUTHOR]

In this work, a novel organic receptor, CPI [(E)-3-(4-(9H-carbazol-9-yl)phenyl)-2-(1H-benzo[d]imidazol-2-yl)acrylonitrile], was rationally designed and successfully fabricated for selective and sole recognition of CN− ions over other competitive anions through an obvious chromogenic and ratiometric emission change in DMSO. The distinct and prominent color change upon the addition of CN− can be attributed to the typical ICT process, which is induced by the deprotonation of acidic NH protons in the imidazole moiety. The sensor displayed strong solvatochromic effects in commonly used organic solvents such as n-hexane, toluene, diethyl ether, DCM, THF, DMF and DMSO. The chemical structure of the sensor was characterized by single-crystal X-ray diffraction, 1HNMR, 13CNMR, IR and mass spectroscopy. Significantly, the probe can function as a fluorescence-based sensor for the efficient detection of low-level water in organic solvents. The solid-state emission properties of CPI were successfully applied to recognise cyanide in a solid-state platform with naked eye-visualized distinct color change. The probe can be made reusable by adding TFA into the CN− treated probe solution. The detection limit of CPI towards CN− was determined to be 4.48 × 10−8 M. More importantly, the sensor is capable of detecting CN− in food samples and has been employed for wastewater treatment. Besides, easy-to-prepare CPI-coated test strips provide a simple, reusable and easy-to-handle protocol for the qualitative identification of CN− conveniently. Finally, density functional theory and time-dependent density functional theory were performed to verify the experimental outcomes theoretically. [ABSTRACT FROM AUTHOR]

Theory of multistep excitation energy migration within the set of chemically identical chromophores distributed on the surface of a spherical nanoparticle is presented. The Green function solution to the master equation is expanded as a diagrammatic series. Topological reduction of the series leads to the expression for emission anisotropy decay. The solution obtained behaves very well over the whole time range and it remains accurate even for a high number of the attached chromophores. Emission anisotropy decay depends strongly not only on the number of fluorophores linked to the spherical nanoparticle but also on the ratio of critical radius to spherical nanoparticle radius, which may be crucial for optimal design of antenna-like fluorescent nanostructures. The results for mean squared excitation displacement are provided as well. Excellent quantitative agreement between the theoretical model and Monte–Carlo simulation results was found. The current model shows clear advantage over previously elaborated approach based on the Padé approximant. [ABSTRACT FROM AUTHOR]

Developing fluorogenic probes for simultaneous live cell labeling of multiple targets is crucial for understanding complex cellular events. The emerging [4+1] cycloaddition between tetrazine and isonitriles holds promise as a bioorthogonal tool, yet existing tetrazine probes lack reactivity and fluorogenicity. Here, we present the development of a series of tetrazine‐functionalized bioorthogonal probes. By incorporating pyrazole adducts into the fluorophore scaffolds, the post‐reacted probes displayed remarkable fluorescence turn‐on ratios, up to 3184‐fold. Moreover, these modifications are generalizable to various fluorophores, enabling a broad emission range from 473 to 659 nm. Quantum chemical calculations further elucidate the turn‐on mechanisms. These probes enable the simultaneous labeling of multiple targets in live cells, without the need for a washing step. Consequently, our findings pave the way for advanced multiplex imaging and detection techniques for cellular studies. [ABSTRACT FROM AUTHOR]

Significance: Breast-conserving surgery (BCS) is limited by high rates of positive margins and re-operative interventions. Fluorescence-guided surgery seeks to detect the entire lesion in real time, thus guiding the surgeons to remove all the tumor at the index procedure. Aim: Our aim was to identify the optimal combination of a camera system and fluorophore for fluorescence-guided BCS. Approach: A systematic review of medical databases using the terms "fluorescence," "breast cancer," "surgery," and "fluorescence imaging" was performed. Cameras were compared using the ratio between the fluorescent signal from the tumor compared to background fluorescence, as well as diagnostic accuracy measures, such as sensitivity, specificity, and positive predictive value. Results: Twenty-one studies identified 14 camera systems using nine different flu-orophores. Twelve cameras worked in the infrared spectrum. Ten studies reported on the difference in strength of the fluorescence signal between cancer and normal tissue, with results ranging from 1.72 to 4.7. In addition, nine studies reported on whether any tumor remained in the resection cavity (5.4% to 32.5%). To date, only three studies used the fluorescent signal for guidance during real BCS. Diagnostic accuracy ranged from 63% to 98% sensitivity, 32% to 97% specificity, and 75% to 100% positive predictive value. Conclusion: In this systematic review, all the studies reported a clinically significant difference in signal between the tumor and normal tissue using various camera/ fluorophore combinations. However, given the heterogeneity in protocols, including camera setup, fluorophore studied, data acquisition, and reporting structure, it was impossible to determine the optimal camera and fluorophore combination for use in BCS. It would be beneficial to develop a standardized reporting structure using similar metrics to provide necessary data for a comparison between camera systems. [ABSTRACT FROM AUTHOR]

Control of the intermolecular aggregation of organic π‐conjugated molecules as chromophores is crucial for tuning their physical properties such as light absorption/emission, and energy and charge transfer. Lots of advances have been achieved in control of intermolecular aggregation of organic chromophores in solid states where an indefinitely large number of molecules are involved. However, much less understanding has been gained at a mesoscale of aggregates formed by well‐defined organization of a deterministic number of chromophores, which has been realized in natural photosynthetic systems but still remains rare in manmade materials. Here, we report both the kinetic and the thermodynamic control of the supramolecular aggregation of a near‐infrared cyanine dye, PPcy, and its derivatives confined in colloidal nanoparticles stabilized by surfactants in aqueous media. Our results demonstrate that both the aggregation number, the aggregation state and the optical properties of the PPcy chromophores are controllable through optimization of the alkyl and polymer chains tethered from PPcy, the effective concentration of the chromophore inside each particle, and the surfactants utilized to stabilize the colloids in water. [ABSTRACT FROM AUTHOR]

Simple Summary: Due to their specificity, monoclonal antibodies have significantly impacted cancer patients' care, becoming one of the fastest-growing classes of new drugs approved for the treatment of solid tumors. Targeted fluorescence-guided surgery is a novel technology to better visualize tumor residuals intraoperatively. It consists of a fluorescent molecular probe, that, once injected, lights up the neoplastic cells during the surgical resection. In this regard, the development of an off-the-shelf large-scale production of clinically approved, fluorescently labeled monoclonal antibodies for targeted fluorescence-guided surgery is becoming an urgent need for oncological surgeons working in this field. Our paper aims to present the current evidence on the clinical use of monoclonal antibodies targeting solid adult and pediatric cancers. Particularly, we aim to define the utility, indications, doses, and timing of administration of the most promising monoclonal antibodies to be used for targeted fluorescence-guided surgery in oncology patients. We will also define the top three monoclonal antibodies that could be used for targeted fluorescence-guided surgery in different solid cancers, to set the basis of a bank of monoclonal antibodies that can be used to deliver highly individualized and personalized surgical approaches. This study aims to review the status of the clinical use of monoclonal antibodies (mAbs) that have completed or are in ongoing clinical trials for targeted fluorescence-guided surgery (T-FGS) for the intraoperative identification of the tumor margins of extra-hematological solid tumors. For each of them, the targeted antigen, the mAb generic/commercial name and format, and clinical indications are presented, together with utility, doses, and the timing of administration. Based on the current scientific evidence in humans, the top three mAbs that could be prepared in a GMP-compliant bank ready to be delivered for surgical purposes are proposed to speed up the translation to the operating room and produce a few readily available "off-the-shelf" injectable fluorescent probes for safer and more effective solid tumor resection. [ABSTRACT FROM AUTHOR]

Labeling and detection of biomolecules in vitro and in vivo is essential to many areas of biomedical science. Fluorophores stand as indispensable tools within chemical biology, underscoring the importance of fine‐tuning their optical properties. This review focuses on methods for optimizing emission wavelength, quantum yield and photostability. We focus not just on the trends, but the fundamental physical organic chemistry concepts that inform the connection between molecular structure and fluorescent properties. This approach offers an essential understanding of fluorescence, enabling readers to develop a systematic analytical framework for thinking about fluorescence. Furthermore, an evaluation of newer non‐planar fluorophores shines light on the bright future of fluorescent molecules. [ABSTRACT FROM AUTHOR]

Organic fluorophores with dual-state emission (DSE) are rare or difficult to observe because most of them display either aggregation-induced emission (AIE) or aggregation-caused quenching (ACQ). Amazing works have been accomplished, yet most of the DSE compounds were excited by UV light which limits their wide application in bioimaging. In this work, we achieved a visible-light excited DSE fluorophore and realized its imaging in SKOV-3 cells and zebrafish. The naphtho[2',3':4,5]imidazo[1,2-a]pyridine (NIP) core ensures its emission in dilute solution. Meanwhile, the twisted phenyl ring blocks fluorescence quenching induced by the π-π stacking and leads to the emission of the solid. The fluorescence intensity is steady even after 6 h of continuous intense sunlight. More importantly, photostability of NIP in cells is much better than commercial dye (mitochondrial green). [ABSTRACT FROM AUTHOR]

RNAs play important roles in regulating biological growth and development. Advancements in RNA-imaging techniques are expanding our understanding of their function. Several common RNA-labeling methods in plants have pros and cons. Simultaneously, plants' spontaneously fluorescent substances interfere with the effectiveness of RNA bioimaging. New technologies need to be introduced into plant RNA luminescence. Aggregation-induced emission luminogens (AIEgens), due to their luminescent properties, tunable molecular size, high fluorescence intensity, good photostability, and low cell toxicity, have been widely applied in the animal and medical fields. The application of this technology in plants is still at an early stage. The development of AIEgens provides more options for RNA labeling. Click chemistry provides ideas for modifying AIEgens into RNA molecules. The CRISPR/Cas13a-mediated targeting system provides a guarantee of precise RNA modification. The liquid–liquid phase separation in plant cells creates conditions for the enrichment and luminescence of AIEgens. The only thing that needs to be looked for is a specific enzyme that uses AIEgens as a substrate and modifies AIEgens onto target RNA via a click chemical reaction. With the development and progress of artificial intelligence and synthetic biology, it may soon be possible to artificially synthesize or discover such an enzyme. [ABSTRACT FROM AUTHOR]

Milk is subjected to different industrial processes, provoking significant physicochemical modifications that impact milk's functional properties. As a rapid and in-line method, front-face fluorescence can be used to characterize milk instead of conventional analytical tests. However, when applying fluorescence spectroscopy for any application, it is not always necessary to determine which compound is responsible for each fluorescent response. In complex matrixes such as milk where several variables are interdependent, the unique identification of compounds can be challenging. Thus, few efforts have been made on the chemical characterization of milk' fluorescent spectrum and the current information is dispersed. This review aims to organize research findings by dividing the milk spectra into areas and concatenating each area with at least one fluorophore. Designations are discussed by providing specific information on the fluorescent properties of each compound. In addition, a summary table of all fluorophores and references cited in this work by area is provided. This review provides a solid foundation for further research and could serve as a central reference. [ABSTRACT FROM AUTHOR]

Temperature is an essential physical characteristic that influences all biological processes. Building on previous research on dialkylamino-functionalized rhodamine-based thermo-sensors, we investigate herein the thermosensitive properties of triamino-phenazinium dyes. Through a simple five-step synthetic route, we synthesized amino-phenazinium chromophores 6 and 7, featuring diethylamine substituents at different positions. A comparative analysis of optical properties and thermosensitivity was conducted on these compounds and an isomer, 5, in which butylamine moiety replaced the diethylamine group. The different emissive behaviors of the three fluorophores emphasize that not only the chemical nature but also the specific position of the alkylamine substituent play fundamental roles in the synthesis of highly emissive thermo-probes.

Fluorescence microscopy remains one of the single most widely applied experimental approaches in neuroscience and beyond and is continuously evolving to make it easier and more versatile. The success of the approach is based on synergistic developments in imaging technologies and fluorophore labeling strategies that have allowed it to greatly diversify and be used across preparations for addressing structure as well as function. Yet, while targeted labeling strategies are a key strength of fluorescence microscopy, they reciprocally impose general limitations on the possible types of experiments and analyses. One recent development that overcomes some of these limitations is fluorescence microscopy shadow imaging, where membranebound cellular structures remain unlabeled while the surrounding extracellular space is made to fluoresce to provide a negative contrast shadow image. When based on super-resolution STED microscopy, the technique in effect provides a positive image of the extracellular space geometry and entire neuropil in the field of view. Other noteworthy advantages include the near elimination of the adverse effects of photobleaching and toxicity in live imaging, exhaustive and homogeneous labeling across the preparation, and the ability to apply and adjust the label intensity on the fly. Shadow imaging is gaining popularity and has been applied on its own or combined with conventional positive labeling to visualize cells and synaptic proteins in their parenchymal context. Here, we highlight the inherent limitations of fluorescence microscopy and conventional labeling and contrast these against the pros and cons of recent shadow imaging approaches. Our aim is to describe the brief history and current trajectory of the shadow imaging technique in the neuroscience field, and to draw attention to its ease of application and versatility. [ABSTRACT FROM AUTHOR]

Fluorophores are considered powerful tools for not only enabling the visualization of cell structures, substructures, and biological processes, but also making for the quantitative and qualitative measurement of various analytes in living systems. However, most fluorophores do not meet the diverse requirements for biological applications in terms of their photophysical and biological properties. Hybridization is an important strategy in molecular engineering that provides fluorophores with complementarity and multifunctionality. This review summarizes the basic strategies of hybridization with four classes of fluorophores, including xanthene, cyanine, coumarin, and BODIPY with a focus on their structure‐property relationship (SPR) and biological applications. This review aims to provide rational hybrid ideas for expanding the reservoir of knowledge regarding fluorophores and promoting the development of newly produced fluorophores for applications in the field of life sciences. [ABSTRACT FROM AUTHOR]

Reactive oxygen and nitrogen species (RONS) are a range of chemical individuals produced by living cells that contribute to the proper functioning of organisms. Cells under oxidative and nitrative stress show excessive production of RONS (including hydrogen peroxide, H 2 O 2 , hypochlorous acid, HOCl, and peroxynitrite, ONOO−) which may result in a damage proteins, lipids, and genetic material. Thus, the development of probes for in vivo detection of such oxidants is an active area of research, focusing on molecular redox sensors, including boronate-caged fluorophores. Here, we report a boronate-based styryl probe with a cationic pyridinium moiety (BANEP+) for the fluorescent detection of selected biological oxidants in vitro and in vivo. We compare the chemical reactivity of the BANEP+ probe toward H 2 O 2 , HOCl, and ONOO− and examine the influence of the major intracellular non-enzymatic antioxidant molecule, glutathione (GSH). We demonstrate that, at the physiologically relevant GSH concentration, the BANEP+ probe is efficiently oxidized by peroxynitrite, forming its phenolic derivative HNEP+. GSH does not affect the fluorescence properties of the BANEP+ and HNEP+ dyes. Finally, we report the identification of a novel type of molecular marker, with the boronate moiety replaced by the iodine atom, formed from the probe in the presence of HOCl and iodide anion. We conclude that the reported chemical reactivity and structural features of the BANEP+ probe may be a basis for the development of new red fluorescent probes for in vitro and in vivo detection of ONOO−. [Display omitted] • Pyridinium-linked styryl-based boronate fluorescent probe for detection of ONOO−. • Iodide ion influences reaction between boronate probes and biological oxidants. • Iodinated dye as a marker for boronate probe and HOCl reaction in the presence of I−. • Glutathione may change products formed in reaction of boronate probe with ONOO−. [ABSTRACT FROM AUTHOR]

We report a rapid, efficient, and scope‐extensive approach for the late‐stage electrochemical diselenation of BODIPYs. Photophysical analyses reveal red‐shifted absorption – corroborated by TD‐DFT and DLPNO‐STEOM‐CCSD computations – and color‐tunable emission with large Stokes shifts in the selenium‐containing derivatives compared to their precursors. In addition, due to the presence of the heavy Se atoms, competitive ISC generates triplet states which sensitize 1O2 and display phosphorescence in PMMA films at RT and in a frozen glass matrix at 77 K. Importantly, the selenium‐containing BODIPYs demonstrate the ability to selectively stain lipid droplets, exhibiting distinct fluorescence in both green and red channels. This work highlights the potential of electrochemistry as an efficient method for synthesizing unique emission‐tunable fluorophores with broad‐ranging applications in bioimaging and related fields. [ABSTRACT FROM AUTHOR]

We report a rapid, efficient, and scope‐extensive approach for the late‐stage electrochemical diselenation of BODIPYs. Photophysical analyses reveal red‐shifted absorption – corroborated by TD‐DFT and DLPNO‐STEOM‐CCSD computations – and color‐tunable emission with large Stokes shifts in the selenium‐containing derivatives compared to their precursors. In addition, due to the presence of the heavy Se atoms, competitive ISC generates triplet states which sensitize 1O2 and display phosphorescence in PMMA films at RT and in a frozen glass matrix at 77 K. Importantly, the selenium‐containing BODIPYs demonstrate the ability to selectively stain lipid droplets, exhibiting distinct fluorescence in both green and red channels. This work highlights the potential of electrochemistry as an efficient method for synthesizing unique emission‐tunable fluorophores with broad‐ranging applications in bioimaging and related fields. [ABSTRACT FROM AUTHOR]

Photoactivatable fluorophores are useful tools in live cell imaging due to precise spatial and temporal control. Herein, a tandem visible‐light photodecaging and tautomerization strategy is proposed as an activation mechanism for the construction of photoconvertible boron dipyrromethenes (BODIPY) dyes, from which nitro‐benzyloxy BODIPY derivatives are developed as a new class of visible light photoactivated fluorophores. Upon exposure to visible light, the nitro‐benzyloxy moiety is released and the BODIPY moiety is converted to the keto‐form BODIPY analog by tautomerization. The photoconversion process not only achieves over 50 nm blue shift of absorption/emission maxima but also provides bright fluorescence for both the pre‐ and post‐activation forms. Mechanism studies indicate that no indications of radical species and singlet oxygen are involved in this unique photoconversion and both UV and visible light irradiation can efficiently activate this photoinduced release of the nitrobenzyl alcohol through photoexciting of BODIPY core rather than classical nitro‐benzyloxy photocage. Cell and vesicle imaging is successfully performed using the new photoconversion dyes. The results show that these new photoconvertible dyes are highly efficient, universal, and non‐cytotoxic, which are valuable for the development of promising photoactivated molecules. [ABSTRACT FROM AUTHOR]

We present the first unambiguous experimental method enabling single-fluorophore sensitivity in a flow cytometer using quantum properties of single-photon emitters. We use a quantum measurement based on the second-order coherence function to prove that the optical signal is produced by individual biomarkers traversing the interrogation volume of the flow cytometer from the first principles. This observation enables the use of the quantum toolbox for rapid detection, enumeration, and sorting of single fluorophores in large cell populations as well as a 'photons-to-moles' calibration of this measurement modality. [ABSTRACT FROM AUTHOR]