Fluorescence Excitation Research Articles

High fluorescence quantum yield of methionine-doped carbon quantum dots for achieving rapid assay of tetracyclines in foodstuffs

The trace-level detection of tetracyclines (TCs) in food products is essential to ensure food safety and public health. Herein, we prepared the methionine-doped carbon quantum dots (Met-CQDs) using citric acid as the precursor. Met-CQDs exhibited a Gaussian unimodal peak centered at 440 nm in the fluorescent excitation spectrum, along with a remarkable greenish-blue emission and a fluorescent quantum yield of 33.5 %. Furthermore, the presence of TC (the quencher) caused a rapid quenching of the fluorescence of Met-CQDs, accompanying with a color transition from light blue to dark bule as TC concentrations increased. The coloring variation was also detected by the images captured by smartphones and RGB analysis software, facilitating portable detection of TC utilizing Met-CQDs as a fluoroprobe. The findings indicate that the Met-CQDs based fluoroprobe exhibits high selectivity, rapid response (only ∼1 min) according to an “ON-OFF” sensing model. This fluorescence sensing method gave a low detection limit (LOD) of 0.032 μM and excellent linearity for TC in the concentration range of 0.1–500 μM. Also, the smartphone-based fluorescence-visualizing approach displayed good linearity with a LOD of 0.33 μM. The interactions between this fluoroprobe and TC occurred by virtue of both inner filter effect (IFE) and static-quenching principle. The average recovery for TC in the milk, honey, and tap water samples was determined to be 98.46 ± 1.71 % by a fluorometric method. Overall, both fluorometric and RGB approaches demonstrate strong correlation with conventional LC-MS/MS, and thus the as-fabricated Met-CQDs are promising for the preliminary screening of TCs’ residues in food products.

Utility of AI digital pathology as an aid for pathologists scoring fibrosis in MASH

Background & AimsIntra and inter-pathologist variability poses a significant challenge in metabolic dysfunction-associated steatohepatitis (MASH) biopsy evaluation, leading to suboptimal selection of patients and confounded assessment of histological response in clinical trials. We evaluated the utility of an artificial intelligence (AI) digital pathology (DP) platform to aid pathologists improve the reliability of fibrosis staging. MethodsA total of 120 digitized histology slides from two trials (NCT03517540, NCT03912532) were analysed by four expert hepatopathologists, with and without AI-assistance in a randomized, cross-over design. We utilized the HistoIndex AI DP platform, consisting of unstained second harmonic generation/two photon excitation fluorescence (SHG/TPEF) images and AI quantitative fibrosis (qF) values. ResultsAI-assistance significantly improved inter-pathologist kappa for fibrosis (F)-staging, particularly for early fibrosis (F0-F2), with reduced variance around the median reads. Intra-pathologist kappa was unchanged. AI-assistance increased pathologist concordance for identifying clinical trial inclusion subjects (F2-F3) from 45% to 71%, exclusion subjects (F0/F1/F4) from 38% to 55%, and evaluation of fibrosis response to treatment from 49% to 61%. SHG/TPEF images, qFibrosis continuous values, and qF-stage were considered useful by at least 3 out of 4 pathologists in 83%, 55%, and 38% cases, respectively. In the context of a clinical trial, the increase in inter-pathologist concordance in this study is modeled to result in a ∼25% reduction in the potential need for adjudication as well as a ∼50% increase in the study power. ConclusionsThe use of AI DP enhances inter-rater reliability of fibrosis staging for MASH. This indicates that the SHG/TPEF-based AI DP tool is useful for assisting pathologists in assessing fibrosis, thereby enhancing clinical trial efficiency and reliability of fibrosis readouts in response to treatments.

Fused-fiber-photometry based on a low-autofluorescence wideband multimode circulator

Analysis of fluorescent signals in biological samples is widely used in the fields of life science and medicine and can be achieved through conceptually distinct approaches. Here, we report the use of a wideband multimode circulator (WMC), a three-port optical fused-fiber device with an asymmetrical structure specifically designed to ensure efficient light transmission in both illumination and collection paths. It features a flat spectral response and is ideally suited for simultaneous excitation and detection of fluorescence in biological tissue. The WMC used in this work was further improved by minimizing its inherent background autofluorescence. Using the WMC, we demonstrate improved fluorescence detection against already existing fiber-based approaches. Finally, we utilize the WMC to establish fiber photometry recordings in vivo to infer neuronal activity in a small neuromodulatory nucleus in the brainstem of an awake mouse.

Excitation-Dependent pKa Extends the Sensing Range of Fluorescence Lifetime pH Sensors

Biological activity is strongly dependent on pH, which fluctuates within a variety of neutral, alkaline, and acidic local environments. The heterogeneity of tissue and subcellular pH has driven the development of sensors with different pKa values, and a huge assortment of fluorescent sensors have been created to measure and visualize pH in living cells and tissues. In particular, sensors that report based on fluorescence lifetime are advantageous for quantitation. Here, we apply a theoretical framework to derive how the apparent pKa of lifetime-based pH sensors depends on fluorescence excitation wavelength. We demonstrate that theory predicts the behavior of two different fluorescent protein-based pH sensors in solution as proofs-of-concept. Furthermore, we show that this behavior has great practical value in living cells because it extends the sensing range of a single sensor by simply choosing appropriate detection parameters to match the physiological pH range of interest. More broadly, our results show that the versatility of a single lifetime-based sensor has been significantly underappreciated, and our approach provides a means to use a single sensor across a range of pH environments.

Blue Light Emitting Electron-Rich Triphenylamine Decorated Stilbene Derivatives for the Bio-Imaging Application

The stilbene functionalized star and bent shaped π-conjugate organic donor-donor (D-π-D) molecules with decorated triphenylamine central core were synthesized via Michaelis-Arbuzov and Wittig-Horner Reaction with excellent regio-specific. The photo-physical investigations clearly indicate the synthesized molecules have excellent optical properties with UV-visible regions and hypochromic shifts. The intra-molecular charge transfer (ICT) absorption of bent and star-shaped molecules was found in the region of 369–390 and 382–390 nm. The star and bent-shaped molecules exhibited emission maximums between 444–490 and 437–476 nm. While increasing the solvent polarity, the emission maximum of TDMSPA and DMSNDMPA molecules were observed in red-shift. The excited state dipole moment changed when compared to ground state due to the triphenylamine core confirmed by the Lippert-Mataga plot. The density functional theory is good evidence; the TDMSPA molecule has a twisted configuration and supports optical and electrochemical studies. Time-correlated single photon counting (TCSPC) technique showed that the excited state fluorescence lifetime of the stilbene molecules was found to be 2.25 and 2.18 ns for star and bent shaped molecules, respectively. The synthesized molecules exhibit significantly low band-gap energies between HOMO and LUMO, and the values are observed in the range between 2.86 and 2.91 eV. The triphenylamine core-based π-conjugate organic chromophore is widely used as a bio-imaging agent in Rhizoctonia oryzae (Plant fungal pathogen) as the model and the TDMSPA was an excellent bio-imaging agent and biocompatibility with fungal and bacterial cells.

Reactive Stroma and Acinar Morphology in Prostate Cancer: Implications for Progression and Prognostic Assessment.

Prostate cancer (PC) remains a significant global health concern, with prognostic assessments largely reliant on the Gleason Classification System. While it has proven effective, subjectivity in interpretation persists, prompting the need for complementary approaches. Reactive stroma (RS) has emerged as a potential candidate for enhancing PC characterization, as it reflects intricate interactions among stromal, epithelial, and extracellular matrix components. To shed light on this, we conducted a comprehensive study. Two expert pathologists independently analyzed consecutive prostate biopsies (n = 120 patients), categorized into four groups based on Gleason scores. Four acinar patterns were described, denoted as A, B, C, and D. Our study uncovered a noteworthy presence of RS, predominantly within poorly differentiated tumors. Stromogenic tumors, characterized by high RS content, were particularly associated with Gleason scores of 4 + 3 and ≥ 8. Intriguingly, acinar patterns, including the distinctive B and D patterns, exhibited strong correlations with stromogenic tumors. Incorporating quantitative imaging techniques (Second Harmonic Generation and Two-Photon Excitation Fluorescence Microscopy), we examined collagen fiber density within the stroma. Our analysis revealed a direct relationship between RS intensity and collagen fiber counts, particularly prominent in patterns B and D. These findings suggest that the stromal reaction in PC is closely linked to acinar morphology and collagen deposition. Moreover, rudimentary microacini at the tumor periphery, associated with intense RS and patterns B and D, may signify an unfavorable prognosis. Our study highlights the potential of RS as an additional prognostic factor in PC. It underscores the intricate interplay between acinar patterns, RS intensity, and collagen fiber density, providing valuable insights for future prognostic assessments and therapeutic strategies. Further exploration of these relationships is essential for a comprehensive understanding of PC progression and management.

Label-Free Detection of Breast Phyllodes Tumors Based onMultiphoton Microscopy.

Phyllodes tumors (PTs) are rare breast stroma neoplasms, and their accurate identification at various stages is essential for personalized patient treatment. In this study, multiphoton microscopy (MPM) with two-photon excited fluorescence (TPEF) and second harmonic generation (SHG) imaging was used for label-free detection and differentiation of PTs and normal breast tissue. An automated image processing strategy was developed to quantify changes in collagen fiber morphology within the stroma and boundary of PTs, establishing optical diagnostic characteristics of PTs using MPM. The results demonstrated that MPM could be used for the detection of different stages of PTs, and the morphological alterations in collagen fibers could serve as critical indicators of PT malignancy, offering new insights for the diagnosis and grading of benign, borderline, and malignant PTs. It lays the groundwork for the future application of compact MPM for the rapid detection and diagnosis of PTs.

Optical Characteristics and Source Analysis of Dissolved Organic Matter in Hypersaline Lakes in Cold and Arid Regions

The composition and source information of the aquatic dissolved organic matter （DOM） in the Daihai Lake, a typical saline lake at the ecologically fragile agro-pastoral area, were explored with three-dimensional excitation and emission matrix fluorescence （3DEEM）, parallel factor analysis （PARAFAC）, and fluorescence peaks （B, T, A, M, C, D, and N）. Further, the relationship between aquatic DOM fluorescence characteristics and environmental factors were analyzed using correlation analysis in the Daihai Lake. The results showed that the water of Daihai was weakly alkaline, with high nutrient salt concentration and serious organic pollution. Two protein-like substances （C1 and C3） and one humic-like substance （C2） were identified using PARAFAC. The sum of C1 + C2 accounted for 80% of DOM. The results of fluorescence peaks showed that the total percentage of protein-like fluorescence absorption peaks （B, T, and N） was 49.67%. Aquatic DOM of Daihai demonstrated both terrigenous and endogenous characteristics. Terrestrial inputs were mainly from intensive agricultural activities and rapid ecological degradation of the watershed leading to large amounts of organic matter entering the lake. The environmental behavior of DOM was highly correlated with the metabolic activity of organisms and nutrient status of lakes. NH4+-N, DOP, TP, DO, and permanganate index characterized the effect of the lake trophic state on DOM composition. DO, SAL, and Chla were mainly related to microbial metabolic processes （P<0.05）. This study provides basic data and insights for the improvement of inland lake environments in saline lakes.

Absorption, Fluorescence, and Two-Photon Excitation Ability of 5-o-Tolyl-11 (or 13)-o-tolylisoindolo[2,1-a]quinolines Prepared by Ring-Closing Metathesis and [2+3] Cycloaddition.

We have successfully improved the fluorescence quantum yield of isoindolo[2,1-a]quinoline derivatives by suppressing the rotation of the phenyl groups at positions 5 and 11 (or 13). Additionally, we found that the planarity of these phenyl groups at positions 5 and 13 of isoindolo[2,1-a]quinoline derivatives is crucial for two-photon absorption properties.

Green hydrothermal approach for the synthesis of carbon quantum dots from waste tea bags for acrylamide detection in drinking water: A fluorescence assay validated by HPLC-PDA analysis

The study focused on converting tea bag waste into strong fluorescence carbon quantum dots (TBW-CQDs) for the detection of acrylamide in drinking water, antimicrobial activity, and photocatalytic degradation. The TBW-CQDs exhibited blue luminescence and maximum absorbance at 287 nm under UV light and distinctive fluorescence emission and excitation wavelengths at 425 nm and 287 nm, respectively. TBW-CQDs revealed a particle size of 8.12 ± 0.06 nm with a spherical morphology followed by an abundance of 59.29 % carbon and 39.82 % oxygen. For acrylamide extraction from water, the QuEChERS method was established, which exhibited a recovery rate of 97 to 99 %. The fluorescence-based sensor exhibited a low limit of detection of 0.35376 ppm, which was validated by HPLC-PDA (LOD 0.300688 ppm). TBW-CQDs degraded 90.62 % of indigo carmine and 93.19 % of methylene blue under bright sunlight. In conclusion, the fabricated TBW-CQDs provide a promising, cost-effective, and precise approach to acrylamide detection in drinking water.

Biofilm growth Dynamics in a Micro-Irrigation with Reclaimed Wastewater in the Field scale.

The dripper clogging due to the development of biofilm can reduce the benefits of micro-irrigation technology implementation using reclaimed wastewater. The narrow cross-section and labyrinth geometry of the dripper channel enhance the fouling mechanisms. The aim of this study was to evaluate the water distribution and biofouling of drip irrigation systems at the field scale during irrigation with treated wastewater. Six 100 m lines of commercial pipes with two pressure-compensating dripper types (flow rate, Q, of 0.65 L.h-1 and 1.5 L.h-1, respectively) were monitored for four months. Different zones along the pipes were selected to evaluate the influence of hydrodynamical conditions (Reynolds number = 5400 to 0) on biofouling. Destructive methods involving the biofilm extraction by mechanical means, showed little biofilm development without significant differences in dry and organic matter content in function of the sampling location along the pipe or dripper flow rate (Q0.65 and Q1.5). These results were confirmed by non-destructive methods, such as optical coherence tomography, that nevertheless showed that biofouling concerned 15-20% of the total dripper labyrinth volume. Total organic carbon monitoring and its composition (by three-dimensional excitation and emission matrix fluorescence microscopy) showed that the biofilm did not significantly influence the organic matter nature. Our results indicated that the biological activity and biofilm development in irrigation systems were more affected by the environmental conditions, particularly water temperature, rather than flow conditions. This confirmed that treated wastewater with low organic content can be used in micro-irrigation systems without significant loss of efficiency, even in conditions requiring intensive irrigation, such as the Mediterranean climate.

Boron-doped carbon dots with time-resolved room temperature phosphorescence for detection of ciprofloxacin hydrochloride and information encryption applications.

Novel boron-doped carbon dots (BCDs) with extended afterglow characteristics were synthesized via a one-step solvothermal method using acrylamide, sulfosalicylic acid, and sodium tetraborate as protective matrices. The presence of boron from sodium tetraborate introduced an empty orbital, allowing it to form a more extended conjugated system with adjacent oxygen atoms, thereby lowering the energy level of the lowest unoccupied molecular orbital in the system. The phosphorescence emission of these BCDs exhibits a red shift over time from 450 to 500 nm. These BCDs have been effectively utilized to produce anti-counterfeit phosphorescent powder. Additionally, the BCDs display optimal fluorescence excitation at 330 nm and optimal emission at 420 nm. They demonstrate a detection limit for ciprofloxacin hydrochloride of 37 nM in the 1-100 µM concentration range and 26 nM in the 100-210 µM range. This fluorescence detection is governed by an inner filter effect. Real sample testing further confirms that these BCDs serve as excellent sensors for ciprofloxacin hydrochloride.

Fluorescence scattering under the Stokes–Mueller formalism determines the orientational distribution of fluorophores and the nature of optically active biological proteins

Abstract In the current paper, we represent intrinsic fluorescence anisotropies as four-dimensional normalized Stokes vectors defined by the maximum excitation and emission in the fluorescence process with respect to linear, linear-45 and circular polarizations of light. Depending upon the transition moments for absorption/excitation and emission of fluorophores, eight types of these Stokes vectors can be realized from the Mueller fluorescence matrix of the system. These Stokes vectors probe the orientational distribution of fluorophores and predict the nature of optically active biological proteins, whether laevorotatory or dextrorotatory. The orthogonality relation between the Stokes vectors corresponding to the excitation and emission processes of fluorescence connects the molecular ground and excited states of biological and non-biological systems.

Ionic Hydrogen-Bonded Organic Frameworks with a Two-Photon Synergistic Color Change and Their Information Encryption Applications.

Photochromic hydrogen-bonded organic frameworks (HOFs) can introduce different luminescent functional groups to achieve synergistic controlled multiple color change properties, which are in great demand for diverse information encryption applications. We report in this paper switchable photochromic and photoluminescent dual luminescent functional group HOFs constructed with synergistic effects by N,N'-bis(2-phenylalanine)-1,4,5,8-naphthalenediimine (H2PheNDI) and benzenecarboximidamide 4,4'-azobis(hydrochloride) (AZBH). The crystal powder of iHOF-41 is orange-red in color, which can be changed to black under the irradiation of a 365 nm ultraviolet (UV) light source for 15 min. The photoisomerization rate of the crystal solution under continuous UV irradiation for 5 h was close to 99%. The composite membranes can achieve the properties of photochromism and photoluminescence when they are discolored under 365 nm UV irradiation and, at the same time, excite red bright fluorescence. This work achieves the construction of HOFs based on switching biluminescent functional groups and explores the synergistic mechanism of the photoisomerization process and photochromism as well as its practical application in information encryption.

Electronic Spectroscopy of Ovalene: Reassignment of the S2(B3u)- S0(Ag) Transition.

As part of our experiments to characterize solid para-hydrogen (para-H2) as a matrix host for electronic spectroscopy with potential applications in the ongoing quest for the carriers of the diffuse interstellar bands (DIB), we studied dispersed fluorescence and fluorescence excitation spectra of ovalene (C32H14), a planar polycyclic aromatic hydrocarbon (PAH) of D2h symmetry. Although generally in good agreement with previously reported data for jet-cooled C32H14, our results, in conjunction with quantum-chemical calculations, indicate that the observed spectral progressions are associated with the S2(B3u)-S0(Ag) electronic transition instead of the originally assigned S1-S0 transition for C32H14 in a supersonic jet, and that the previously reported origin band was misassigned and should be located at ∼21050 cm-1. The reassignment is further supported by the comparably long fluorescence lifetime of ∼1.7 μs. From an analysis of spectral features located >1600 cm-1 in the fluorescence excitation spectrum, we estimate an S2(B3u)-S3(B1g) energy gap of ∼350 cm-1.

Evidence for ecological tuning of anuran biofluorescent signals

Although biologists have described biofluorescence in a diversity of taxa, there have been few systematic efforts to document the extent of biofluorescence within a taxonomic group or investigate its general significance. Through a field survey across South America, we discover and document patterns of biofluorescence in tropical amphibians. We more than triple the number of anuran species that have been tested for this trait. We find evidence for ecological tuning (i.e., the specific adaptation of a signal to the environment in which it is received) of the biofluorescent signals. For 56.58% of species tested, the fluorescence excitation peak matches the wavelengths most abundant at twilight, the light environment in which most frogs are active. Additionally, biofluorescence emission spans both wavelengths of low availability in twilight and the peak sensitivity of green-sensitive rods in the anuran eye, likely increasing contrast of this signal for a conspecific receiver. We propose an expanded key for testing the ecological significance of biofluorescence in future studies, providing potential explanations for the other half of fluorescent signals not originally meeting formerly proposed criteria. With evidence of tuning to the ecology and sensory systems of frogs, our results suggest frog biofluorescence is likely functioning in anuran communication.

Complex-valued scatter compensation in nonlinear microscopy

Nonlinear, i.e., multiphoton microscopy is a powerful technique for imaging deep into biological tissues. Its penetration depth can be increased further using adaptive optics. In this work, we present a fast, feedback-based adaptive-optics algorithm, termed C-DASH, for multiphoton imaging through multiple-scattering media. C-DASH utilizes complex-valued light shaping (i.e., joint shaping of amplitude and phase), which offers several advantages over phase-only techniques: it converges faster, it delivers higher image quality enhancement, it shows a robust performance largely insensitive to the axial position of the correction plane, and it has a higher ability to cope with, on the one hand, scattering media that vary over time and, on the other hand, scattering media that are partially absorbing. Furthermore, our method is practically self-aligning. We also present a simple way to implement C-DASH using a single reflection off a phase-only spatial light modulator. We provide a thorough characterization of our method, presenting results of numerical simulations as well as two-photon excited fluorescence imaging experiments. Published by the American Physical Society 2024

Preparation and Characterization of Novel α-Naphthol Luminophors for Optoelectronic Devices.

Luminophors of α-Naphthol doped with varying concentrations of anthracene and perylene were prepared by conventional solid state reaction method to explore its fluorescence characteristics. The fluorescence spectra of bicomponent and tricomponent luminophors reveals the fluorescence excitation energy transfer (FRET) process. The prepared bi and tri component luminophors were excited at 290nm which corresponds to host excitation wavelength. The weak violet fluorescence of host, α-Naphthol, gets quenched and give anthracene like emission in bicomponent luminophor systems. Further doping with second host, perylene, in anthracene containing α-Naphthol luminophors, red shifted emission was observed at 605nm. The structural parameters, thermal stability and electrical properties of doped luminophors were studied by using X-ray diffraction, TGA-DSC and Cyclic Voltammetry respectively. The particle size of the prepared luminophors was confirmed by scanning electron microscopy (SEM).

Application Exploration of the Photon Chip Technology in Real time Processing of Biomedical Images

In the biomedical field, photonic chip technology is gradually becoming the key to improving the speed and accuracy of medical imaging. The development of this technology has given us new hope in early disease diagnosis and cell monitoring. Recent research has not only sorted out the hotspots and future trends of biomedical photonics, but also paid special attention to the application of photon technology in these fields. Specifically, photonic chips have demonstrated their unique capabilities in multi-color two-photon excitation fluorescence microscopy imaging, two-photon fluorescence lifetime imaging, and two-photon fiber endoscopic imaging. These applications not only improve the clarity of imaging, but also accelerate processing speed, which is crucial for rapid diagnosis and treatment. At the end of the article, the author summarized the contributions of photonic chip technology in the field of biomedical imaging and provided prospects for future development directions.

HER4 is a high-affinity dimerization partner for all EGFR/HER/ErbB family proteins.

Human epidermal growth factor receptors (HER)-also known as EGFR or ErbB receptors-are a subfamily of receptor tyrosine kinases (RTKs) that play crucial roles in cell growth, division, and differentiation. HER4 (ErbB4) is the least studied member of this family, partly because its expression is lower in later stages of development. Recent work has suggested that HER4 can play a role in metastasis by regulating cell migration and invasiveness; however, unlike EGFR and HER2, the precise role that HER4 plays in tumorigenesis is still unresolved. Early work on HER family proteins suggested that there are direct interactions between the four members, but to date, there has been no single study of all four receptors in the same cell line with the same biophysical method. Here, we quantitatively measure the degree of association between HER4 and the other HER family proteins in live cells with a time-resolved fluorescence technique called pulsed interleaved excitation fluorescence cross-correlation spectroscopy (PIE-FCCS). PIE-FCCS is sensitive to the oligomerization state of membrane proteins in live cells, while simultaneously measuring single-cell protein expression levels and diffusion coefficients. Our PIE-FCCS results demonstrate that HER4 interacts directly with all HER family members in the cell plasma membrane. The interaction between HER4 and other HER family members intensified in the presence of a HER4-specific ligand. Our work suggests that HER4 is a preferred dimerization partner for all HER family proteins, even in the absence of ligands.