Ratiometric Fluorescence Research Articles

Amine vapor-responsive ratiometric sensing tag based on HPTS/TPB-PVA fluorescent film for visual determination of fish freshness

In this study, amine vapor-sensitive films with ratiometric fluorescence attributes were developed. The pH-sensitive fluorescein 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt (HPTS) and its tetraphenylethylene derivative (TPB) were selected as ratiometric indicators and incorporated into a polyvinyl alcohol (PVA) matrix to produce HPTS/TPB-PVA films. The films responded well to amine vapors, and the interference of aromatic vapors did not substantially affect the fluorescence signals of the films. Under UV light at a wavelength of 365 nm, the fluorescence of the films changed from dark pink to light pink and finally to yellow when the freshness of the fish was visually checked during storage. In addition, the color difference values of the films showed a positive correlation with the total volatile basic nitrogen (TVB-N), ranging from 12.7 to 24.8 mg/100 g at 25 °C and 8.4 to 25.6 mg/100 g at 4 °C, respectively. This indicates that fluorescent films have good potential for quantifying fish freshness in the near future when connected to an automatic data processing system based on color differences.

Synthesis and application of hypochlorite ratiometric fluorescence probe based on cellulose

Cellulose is the most abundant natural polymer with good biocompatibility and easy modification characteristics. In this paper, a novel cellulose fluorescence probe CNS for detecting ClO− was prepared by modifying microcrystalline cellulose (MCC). The fluorescence detection results indicate that CNS exhibits a highly specific “ratiometric” and “colorimetric” fluorescence response to ClO−. In the presence of ClO−, the fluorescence color changes from green to cyan. In addition, the color of the solution changes from yellow to colorless, which can be observed with the “naked eye”. Considering the good selectivity and anti-interference ability of CNS, the probe can be used for the detection of ClO− in real water samples. Importantly, CNS composite films and test papers were prepared and showed practicability in the detection of ClO−, highlighting its broad application potentials.

An intelligent ratiometric fluorescent assay based on MOF nanozyme-mediated tandem catalysis that guided by contrary logic circuit for highly sensitive sarcosine detection and smartphone-based portable sensing application

As the well-known test-indicator for early prostate cancer (PCa), sarcosine (SA) is closely related to the differential pathological process, which makes its accurate determination increasingly significant. Herein, we for the first time expanded the peroxidase (POD)-like property of facile-synthesized Zn-TCPP(Fe) MOF to fluorescent substrates and exploited it to ratiometric fluorescent (RF) sensing. By harnessing the effective catalytic oxidation of MOF nanozyme toward two fluorescent substrates (Scopoletin, SC; Amplex Red, AR) with contrary changes, and target-responsive (SA + SOx)/MOF/(SC + AR) tandem catalytic reaction, we constructed the first MOF nanozyme-based RF sensor for the quantitative determination of SA. Superior to previous works, the operation of this RF sensor is under the guidance of AND-(AND^NAND) contrary logic circuit. The dual-channel binary output changes (from 1/0 to 0/1) not only enable the intelligent logical recognition of SA, bringing strengthened reliability and accuracy, but also manifest the proof-of-concept discrimination of PCa individuals and healthy ones. Through recording the fluorescence alterations of SC (F465) and AR (F585), two segments of linear relationships between ratiometric values (F585/F465) and varied contents of SA are realized successfully. The LOD for SA could reach to as low as 39.98 nM, which outperforms all nanozyme-originated SA sensors reported till now. Moreover, this sensor also demonstrates high selectivity and satisfactory performance in human serum samples. Furthermore, the portable sensing of SA is realized under the assistance of smartphone-based RGB analysis, demonstrating the potential of point-of-care diagnostics of PCa in the future.

Rational design of a near-infrared dual-emission fluorescent probe for ratiometric imaging of glutathione in cells.

Sensors for which the output signal is an intensity change for a single-emission peak are easily disturbed by many factors, such as the stability of the instrument, intensity of the excitation light, and biological background. However, for ratiometric fluorescence sensors, the output signal is a change in the intensity ratio of two or more emission peaks. The fluorescence intensity of these emission peaks is similarly affected by external factors; thus, these sensors have the ability to self-correct, which can greatly improve the accuracy and reliability of the detection results. To accurately image glutathione (GSH) in cells, gold nanoclusters (AuNCs) with intrinsic double emission at wavelengths of 606nm and 794nm were synthesized from chloroauric acid. With the emission peak at 606nm as the recognition signal and the emission peak at 794nm as the reference signal, a near-infrared dual-emission ratio fluorescence sensing platform was constructed to accurately detect changes in the GSH concentration in cells. In vitro and in vivo analyses showed that the ratiometric fluorescent probe specifically detects GSH and enables ultrasensitive imaging, providing a new platform for the accurate detection of active small molecules.

A background correctable fluorescence sensing strategy for adenosine triphosphate evaluation based on aptamer configuration alteration

Ratiometric sensors with background correction are expected to get around the interference from complicated systems. However, there is currently a scarcity of viable carriers for quickly and easily assembling fluorophores with varying fluorescence emission. In this work, we developed a novel aptamer-based ratiometric fluorescence sensor for analysis of adenosine triphosphate (ATP) based on the properties of aptamer that enable conformational transformations upon binding to the target. The carbon dots (CDs) and thioflavin T (ThT) are employed as the reference and detection signal for the ratio-dependent sensor, which are both integrated with the aptamer. The green fluorescence of ThT was initially turn-on after binding with aptamer and then quenched in the presence of ATP, whereas the red fluorescence of CDs remained practically steady. This approach exhibits remarkable selectivity and can detect ATP in the range of 0.5–60 μM with a detection limit of 0.167 μM. Moreover, the method was successfully applied to the determination of ATP in human serum as well as in A549 cells, demonstrating the potential for medical diagnostic applications.

Albumin Hydrogel-Coated Mesoporous Silica Nanoparticle as a Carrier of Cationic Porphyrin and Ratiometric Fluorescence pH Sensor.

Here, we report that a mesoporous silica nanoparticle (MSN) coated with a fluoresceine-labeled bovine serum albumin (F-BSA) hydrogel layer works as a temperature-responsive nanocarrier for tetrakis-N-methylpyridyl porphyrin (TMPyP) and as a fluorescence ratiometric pH probe. F-BSA hydrogel-coated MSN containing TMPyP (F-BSA/MSN/TMPyP) was synthesized by thermal gelation of denatured F-BSA on the external surface of MSN. The F-BSA hydrogel layer was composed of an inner hard corona layer and an outer soft layer and was stable under physiological conditions. F-BSA/MSN/TMPyP exhibited temperature-dependent exponential release of TMPyP. In this release profile, the MSN was found to be a suitable host for stable encapsulation of tetracationic TMPyP by electrostatic interactions, and the F-BSA hydrogel layer mediated the diffusion of TMPyP from the MSN pore interior into the solution phase. Increasing temperature promoted partitioning of TMPyP from the pore interior to the F-BSA hydrogel layer, from where it was spontaneously released into the bulk solution phase by cation exchange. F-BSA/MSN/TMPyP also gave a linear ratiometric fluorescence response (1.3 per pH unit) in the pH range from 6.1 to 8.9.

Silver nanocluster-based ratiometric fluorescence sensors for X-ray dose detection

In this paper, we synthesized silver nanoclusters using bovine serum albumin (BSA) as a template (BSA@AgNCs). Then, we anchored hydroxyphenyl fluorescein (HPF) to yield HPF-BSA@AgNCs. When exposed to X-rays, hydroxyl (∙OH) radicals generated by radiolysis of water react with HPF to produce fluorescein, which emits enhanced fluorescence at 515 nm (λex = 480 nm). The fluorescence intensity of BSA@AgNCs at 685 nm (λex = 480 nm) remains stable when exposed to X-rays. This HPF-BSA@AgNCs ratiometric fluorescence sensor can rapidly detect 0.1–20 Gy (the energy deposited per unit mass, J/kg) of X-rays. In addition, HPF-BSA@AgNCs exhibit good durability and temperature stability. Finally, HPF-BSA@AgNCs were used to measure the absorbed doses of A549 cells and evaluate the cell irradiation damage.

Yolk-shell nanospheres based silane-functionalized carbon dots: Designed synthesis and application in highly selective determination of aflatoxin biosynthesis-related genes nor-1 in cereal crops

This study presented advanced monitoring approaches for the aflatoxin gene cluster in Aspergillus flavus and Aspergillus parasiticus, which are effective tools for controlling aflatoxin proliferation in the food chain. A novel approach involved a ratiometric fluorescence probe based on yolk-shell nanospheres (bCDs@SiO2@gQDs). The probe's fluorescence was activated by DNA hybridization with the modified bCDs@SiO2@gQDs and gold nanoparticles (AuNPs), functioning through donor–acceptor pairs. The exceptional properties of the gQDs and the high FRET efficiency of this system established a functionalized platform for DNA labeling. When the nor-1 gene was present, FRET between the gQDs and AuNPs triggered fluorescence quenching, enabling nor-1 gene detection. The yolk-shell nanostructure of the fluorescent nanoprobes provided an internal reference, reducing experimental errors due to background noise, instrumentation factors, and probe concentration. This allowed for accurate and highly sensitive quantitative determination of the nor-1 gene. The ratiometric fluorescence assay demonstrated a broad linear range (0.1–100 nM) and a low detection limit (12.83 pM) for the nor-1 gene. Tests on non-contaminated maize samples spiked with various concentrations of the nor-1 gene achieved recovery rates between 95.43 % and 101.46 %. This biosensor represents a significant advancement in food safety, offering a simple, rapid, and sensitive method for monitoring agricultural products.

Eu3+-Doped aluminum Metal-Organic Frameworks: A versatile platform for ratiometric detection of histidine and Water, and visible fingerprint identification

Ratiometric fluorescent sensors possess inherent self-calibration features that enable the mitigation of interference from factors unrelated to the analysis. This characteristic results in heightened sensitivity and more pronounced visual detection during qualitative and quantitative analytical processes. A ratiometric fluorescence probe Eu/Al-MOF has been successfully prepared via Eu3+ doped aluminum metal − organic frameworks. Upon introducing histidine, the fluorescence intensity of Eu3+ showed a noteworthy increase, whereas the intensity originating from the ligand decreased owing to enhancing energy transfer from the ligand to Eu3+ ions in the probe. Eu/Al-MOF could detect histidine over a wider linear range of 0.1–400 μM and a lower detection limit of 0.04 μM. Besides, in the presence of water in organic solvents, the excited state of Eu3+ in the probe could be coupled with the O-H oscillation of water molecules, and the excited state energy from Eu3+ would be transferred to the water molecules so that only the fluorescence intensity of Eu3+ gradually decreased. Thus, the proportional detection of trace water could be realized. In ethanol and DMF, the detection range for water was 0.1–20 % and 0.5–20 %, respectively. Moreover, it was found that Eu/Al-MOF could efficiently achieve the visualization identification of fingerprints, providing the possibilities for applying the nano-sensor as a promising multifunctional fluorescence probe.

Detection of Amaranth and Iron in Foods by Modified Carbon Quantum Dots as a Ratiometric Fluorescence Probe

AbstractIn this work, a new carbon quantum dots (CDs) was prepared though microwave assisted approach using citric acid anhydrous (CA) and N‐[3‐(Trimethoxysilyl)propyl] ethylenediamine (TPED) as raw materials. However, the CDs was not effective in actual sensing applications, so further modification of CDs was done by 3,5‐Dichloro‐2‐hydroxybenzaldehyde (it was named as G‐CDs). Fortunately, the modified CDs (G‐CDs) has good selectivity for Fe3+ and amaranth. Studies confirmed that Fe3+ could quench the fluorescence of G‐CDs by forming a complex, while the fluorescence quenching mechanism of G‐CDs by amaranth (AM) mainly belonged to the internal filtering effect (IFE). A detection limit of 20.8 nM for Fe3+ and 43.8 nM for AM were achieved. Moreover, the as‐developed G‐CDs probe was also successfully used to detect Fe3+ and amaranth in real samples, and compared with their results by the standard method, hinting that the accuracy and reliability of the probe were highly satisfactory.

Two birds with one stone: A multi-functional nanoplatform for sensitive detection and real-time inactivation of pathogenic bacteria with NIR-triggered PTT/PDT

Given the increase in public safety incidents due to bacterial infections, sensitive detection and real-time inactivation of pathogenic bacteria have garnered increasing attention in the field of food safety, clinical diagnosis, and environmental monitoring. In this study, we constructed a “two-in-one” PB@DPA-Ce-GMP@Van (PCV) nanoplatform possessing enzyme-like properties and photothermal therapeutic (PTT) and photodynamic therapeutic (PDT) abilities for the detection and inactivation of pathogenic bacteria. PCV was captured on the surface of S. aureus to form a PCV/S. aureus complex. After centrifugation, PCV suspensions could catalyze non-fluorescent Amplex Red (AR) into fluorescent substrates and fluorescent scopolamine (SC) into non-fluorescent substrates. Hence, a ratiometric fluorescence probe with SC/AR reading was constructed for the sensitive detection of bacteria. The detection range reached 101–107 colony-forming unit (CFU)/mL, and the detection limit was as low as 5.0 CFU/mL. Furthermore, approximately 99.7 % of free S. aureus can be inactivated by the PTT and PDT abilities of PCV. Thus, this nanoplatform exhibits a novel advantage in detecting and inactivating bacteria. Moreover, the bacterial infection wound model indicated that PVC exerts good disinfection ability. This study is of immense significance in the application of targeted sensing, eliminating bacterial infection, and even clinical therapy.

An enzyme-free and label-free ratiometric fluorescence signal amplification biosensor based on DNA-silver nanoclusters and catalytic hairpin assembly for tetracycline detection

Sensitive and accurate in antibiotic detection is crucial for food security, environmental conservation, public health, and environmental protection. In this study, we developed an enzyme-free and label-free ratiometric fluorescence biosensor for highly sensitive detection of tetracycline (TET). This biosensor utilized a combination of target-triggered catalytic hairpin assembly (CHA) and DNA-silver nanocluster (DNA-AgNCs) dual-color fluorescent signal probes. Upon TET binding to the aptamer, it initiated the leakage of CHA initiator, resulting in the formation of numerous double-stranded DNA complexes. This process exposed the foothold sequence of hairpin-structured DNA-AgNCs (H3-AgNCs) that exhibit yellow fluorescence. Subsequently, the conformational changes in H3-AgNCs significantly reduced the yellow fluorescence intensity (FI) and reveal the auxiliary DNA-AgNCs (a-DNA-AgNCs) linkage sequence. The binding of a-DNA-AgNCs brought two dark AgNCs close, forming nanocluster dimers emitting red fluorescence. The results demonstrate that the variation in ratiometric signal is linear with TET concentration between 0.1 and 50 ng/mL, with a detection limit as low as 0.086 ng/mL. Additionally, this approach exhibited good recovery results with spiked milk samples. This enzyme-free and label-free combined signal amplification method with proportional signal output based on DNA-AgNCs provides valuable empirical reference for creating low-cost, stable, and highly-sensitive biosensors.

A ternary nucleotide-lanthanide coordination nanoprobe for ratiometric fluorescence detection of ciprofloxacin.

Ciprofloxacin (CIP) is a widely used broad-spectrum antibiotic and has been associated with various side effects, making its accurate detection crucial for patient safety, drug quality compliance, and environmental and food safety. This study presents the development of a ternary nucleotide-lanthanide coordination nanoprobe, GMP-Tb-BDC (GMP: guanosine 5'-monophosphate, BDC: 2-amino-1,4-benzenedicarboxylic acid), for the sensitive and ratiometric detection of CIP. The GMP-Tb-BDC nanoprobe was constructed by incorporating the blue-emissive ligand BDC into the Tb/GMP coordination polymers. Upon the addition of CIP, the fluorescence of terbium ion (Tb3+ ) was significantly enhanced due to the coordination and fluorescence sensitization properties of CIP, while the emission of the BDC ligand remained unchanged. The nanoprobe demonstrated good linearity in the concentration range of 0-10 μM CIP. By leveraging mobile phone software to analyze the color signals, rapid on-site analysis of CIP was achieved. Furthermore, the nanoprobe exhibited accurate analysis of CIP in actual drug and milk samples. This study showcases the potential of the GMP-Tb-BDC nanoprobe for practical applications in CIP detection.

A ratiometric fluorescent probe revealing the abnormality of acetylated tau by visualizing polarity in Alzheimer's disease.

Abnormal neuronal polarity leads to early deficits in Alzheimer's disease (AD) by affecting the function of axons. Precise and rapid evaluation of polarity changes is very important for the early prevention and diagnosis of AD. However, due to the limitations of existing detection methods, the mechanism related to how neuronal polarity changes in AD is unclear. Herein, we reported a ratiometric fluorescent probe characterized by neutral molecule to disclose the polarity changes in nerve cells and the brain of APP/PS1 mice. Cy7-K showed a sensitive and selective ratiometric fluorescence response to polarity. Remarkably, unlike conventional intramolecular charge transfer fluorescent probes, the fluorescence quantum yield of Cy7-K in highly polar solvents is higher than that in low polar solvents due to the transition of neutral quinones to aromatic zwitterions. Using the ratiometric fluorescence imaging, we found that beta-amyloid protein (Aβ) inhibits the expression of histone deacetylase 6, thereby increasing the amount of acetylated Tau protein (AC-Tau) and ultimately enhancing cell polarity. There was a high correlation between polarity and AC-Tau. Furthermore, Cy7-K penetrated the blood-brain barrier to image the polarity of different brain regions and confirmed that APP/PS1 mice had higher polarity than Wild-type mice. The probe Cy7-K will be a promising tool for assessing the progression of AD development by monitoring polarity.

Ratiometric fluorescence detection of Hg2+ based on gold nanocluster/carbon quantum dots nanohybrids.

Ratiometric fluorescence sensing methods are widely used in analysis and detection due to their high sensitivity and stability. In this work, a ratiometric fluorescence method for sensitive detection of Hg2+ was established using a gold nanoclusters/carbon quantum dots (AuNCs/CQDs) nanohybrid probe. The AuNCs/CQDs nanohybrids probe were simply constructed by mixing blue-light-emitting gold nanoclusters (AuNCs) with an orange-emissive carbon quantum dots (CQDs). The probe had two fluorescence emission peaks at 434 nm and 561 nm when the excitation wavelength was 375 nm. With the addition of Hg2+, the fluorescence at 434 nm decreased and the fluorescence at 561 nm remained unchanged; the fluorescence intensity ratio Δ(F434/F561) and Hg2+ concentration have a good linear relationship in the range of 8.32 × 10-7 to 7.69 × 10-5 mol L-1, and the limit of detection (LOD) is 3.58 × 10-7 mol L-1. The method was applied in the detection of Hg2+ in cosmetics and wastewater, and has potential applications for detecting Hg2+ in other samples.

A ratiometric fluorescence assay for the detection of DNA methylation based on an alkaline phosphatase triggered in situ fluorogenic reaction.

The accurate and sensitive quantification of DNA methylation is significant for the early diagnosis of cancer. In this work, an alkaline phosphatase (ALP) triggered in situ fluorogenic reaction between ascorbic acid (AA) and 2,3-DAN was employed as a ratiometric fluorescent probe for the accurate and sensitive detection of DNA methylation with the assistance of ALP encapsulated liposomes. The quinoxaline derivative with a yellow fluorescence emission (I525) was generated from the reaction between AA and 2,3-DAN. Meanwhile, the consumption of 2,3-DAN declined its fluorescence intensity (I386). A ratiometric fluorescent probe (I525/I386) constructed by the above in situ fluorogenic reaction was applied for the accurate detection of DNA methylation. The methylated DNA was first captured by its complementary DNA in 96-well plates. Then, 5mC antibody (Ab) linked liposomes that were encapsulated with ALP recognized and combined with the methylation sites of the target DNA. After the liposomes were lysed by Triton X-100, the released ALP triggered the hydrolysis of ascorbic acid diphosphate (AAP) to form AA, participating in the fluorogenic reaction with 2,3-DAN to produce a quinoxaline derivative. Thus, the ratiometric fluorescence detection of DNA methylation was achieved using I525/I386 values. Using the ALP-enzyme catalyzed reaction and liposomes as signal amplifiers, a low detection limit of 82 fM was obtained for DNA methylation detection. Moreover, the accuracy of the assay could be improved using ratiometric fluorescent probes. We hope that the proposed assay will pave a new way for the accurate determination of low-abundance biomarkers.

Construction of a novel aminofluorene-based ratiometric near-infrared fluorescence probe for detecting carboxylesterase activity in living cells.

Herein, we constructed a novel aminofluorene-based fluorescence probe (FEN-CE) for the detection of carboxylesterase (CE) in living cells by a ratiometric near-infrared (NIR) fluorescence signal. FEN-CE with NIR emission (650 nm) could be hydrolyzed specifically by CE and transformed to FENH with the release of the self-immolative group, which exhibited a red-shifted emission peak of 680 nm. In addition, FEN-CE showed high selectivity for CE and was successfully used in the detection of CE activity in living cells through its ratiometric NIR fluorescence signals.

An efficient ratiometric fluorescence and colorimetric dual-mode probe for convenient determination of nitrite in real samples and E. coli.

Developing an effective and convenient nitrite detection method is indispensable in food safety, environmental monitoring, clinical diagnosis of diseases, and many other areas. Herein, a dicyanoisophorone derivative, TMN-NH2 with large Stokes shift and near-infrared (NIR) emission, was proposed as a ratiometric fluorescence and colorimetric dual-mode probe for the rapid determination of NO2- in acidic media, showing excellent selectivity and high sensitivity. The sensing mechanism is based on the diazotization of TMN-NH2 with NO2- and subsequent diazonium salt hydrolysis to form a hydroxyl-substituted product (TMN-OH). Under the optimized conditions of reaction and detection, a new quantitative analysis method based on TMN-NH2 was established for NO2- detection, exhibiting good linear relationships to NO2- in the range of 0.5 to 15 μM with practical detection limits of 26.6 nM and 17.6 nM for the colorimetric and fluorescent readout, respectively. The quantitative detection of NO2- in real samples demonstrated satisfactory recoveries and repeatability. Moreover, TMN-NH2 was successfully applied for monitoring NO2- in Escherichia coli by confocal fluorescence imaging.

Terbium alginate encapsulated CsPbI3@Pb-MOF: a ratiometric fluorescent bead for detection and adsorption of Fe3.

Halide perovskite nanocrystals are innovative luminescent materials for fluorescent probes with high quantum yield and narrow emission bandwidth. However, the limited stability, single-signal response, and separation challenges obstruct their widespread use in water ion detection. Herein, a ratiometric fluorescence sensor based on terbium alginate gel beads (green fluorescent, namely Tb-AG) embedded with powdered CsPbI3@Pb-MOF (red fluorescent) was prepared for fluorescent determination and adsorption of Fe3+. Pb-MOF's protection notably enhances the water stability of CsPbI3, while the energy transfer between CsPbI3@Pb-MOF and Tb3+ elevates the optical performance of CsPbI3@Pb-MOF@Tb-AG. Significantly, Fe3+ markedly suppresses CsPbI3@Pb-MOF red fluorescence at 647 nm, while not noticeably affecting Tb-AG green emission at 528 nm. The sensor exhibited a strong linear response to Fe3+ concentrations ranging from 0 to 90 μM, with a detection limit of 0.44 μM and high selectivity. The CsPbI3@Pb-MOF@Tb-AG-based sensor has been effectively validated through its successful use in detecting Fe3+ in tap and river water samples. Furthermore, CsPbI3@Pb-MOF@Tb-AG demonstrates a notable adsorption capacity of 325.4 mg g-1 Fe3+. Finally, the mechanism of Fe3+ detection and adsorption was determined.

Pore-coordination sphere facilitated ratiometric fluorescent detection of organic amines in a substitutional metal–organic matrix

A substitutional metal–organic matrix exhibits a highly-sensitive ratiometric fluorescence response towards ammonia and aliphatic amines based on the pore-coordination sphere interaction.