Lysosomes Initiating and DNAzyme-Assisted Intracellular Signal Amplification Strategy for Quantification of Alpha-Fetoprotein in a Single Cell.

Cellular trace proteins are critical for maintaining normal cell functions, with their quantitative analysis in individual cells aiding our understanding of the role of cell proteins in biological processes. This study proposes a strategy for the quantitative analysis of alpha-fetoprotein in single cells, utilizing a lysosome microenvironment initiation and a DNAzyme-assisted intracellular signal amplification technique based on electrophoretic separation. A nanoprobe targeting lysosomes was prepared, facilitating the intracellular signal amplification of alpha-fetoprotein. Following intracellular signal amplification, the levels of alpha-fetoprotein (AFP) in 20 HepG2 hepatoma cells and 20 normal HL-7702 hepatocytes were individually evaluated using microchip electrophoresis with laser-induced fluorescence detection (MCE-LIF). Results demonstrated overexpression of alpha-fetoprotein in hepatocellular carcinoma cells. This strategy represents a novel technique for single-cell protein analysis and holds significant potential as a powerful tool for such analyses.

A Framework Nucleic Acid-Mediated Multimodal Tandem Multivariate Signal Amplification Strategy for Simultaneous Absolute Quantification of Three Different MicroRNAs in a Single Cell.

The simultaneous quantification of multiple microRNAs (miRNA) in a single cell can help scientists understand the relationship between different miRNA groups and different types of cancers from an miRNA omics perspective at the single-cell level. However, there currently remains a challenge in developing techniques for the simultaneous absolute quantification of multiple miRNAs in single cells. Herein, we propose a framework nucleic acid (FNA)-mediated multimodal tandem multivariate signal amplification strategy for simultaneous absolute quantification of three different miRNAs in a single cell. In this study, DNA hexahedron FNAs (DHFs) and DNA tetrahedron FNAs (DTFs) were first prepared, multiple DNA hairpins and substrates were then connected to the hexahedron frame nucleic acid as the target recognition units, and three substrates with labeled FAM fluorophores on the tetrahedral frame nucleic acid served as signal output units. After the two types of FNAs entered the cell, they reacted with three different miRNAs (miRNA-155, miRNA-373, and miRNA-21) and multimodal tandem multivariate signal amplification was initiated simultaneously, reducing the detection limit of the three miRNAs to 8 × 10-15, 2 × 10-15, and 1 × 10-15 M, respectively. The detection sensitivity of the three miRNAs was simultaneously increased by six orders of magnitude, reaching the quantitative requirement of trace miRNAs in single cells. Combined with single-cell injection, membrane melting, and intracellular component separation technology on a microchip electrophoresis platform, we achieved the simultaneous absolute quantification of three different miRNAs in a single cell, thereby providing an important novel method that can be used to conduct single-cell research.

Fluidic Membrane Accelerating the Kinetics of Photoactivatable Hybridization Chain Reaction for Accurate Imaging of Tumor-Derived Exosomes.

Slow intermolecular collisions and "always active" responses compromise the amplification efficiency and response accuracy of nonenzymatic hybridization chain reaction (HCR). In this study, a photoactivatable membrane-oriented HCR (MOHCR) system was rationally designed by binding a photocleavable initiator probe onto a target protein and then anchoring cholesterol-modified hairpin-structure fuel probes. When irradiated, the bound initiator probe was photoactivated and initiated self-assembly to generate activatable and amplified imaging. In a proof-of-concept assay, breast-cancer-derived exosomes were imaged based on the surface protein epithelial cell adhesion molecule (EpCAM). Photoactivatable responses provided precise spatiotemporal control of the MOHCR, and fluidic membranes enabled accelerated reaction kinetics. Our MOHCR system demonstrated high efficiency and accuracy in differentiating between plasma samples from breast cancer patients and healthy donors.

Intracellular Multicomponent Synchronous DNA-Walking Strategy for the Simultaneous Quantification of Tumor-Associated Proteins in a Single Cell.

Single-cell protein analysis is very important for understanding cellular heterogeneity and single-cell biology. However, owing to the extremely low levels of some tumor-associated proteins in individual cells, the absolute quantification of such tumor-associated proteins in a single cell remains a challenge. Herein, an intracellular multicomponent synchronous DNA-walking strategy is proposed for the simultaneous quantification of multiple tumor-associated proteins in a single cell. In this strategy, a nanoprobe based on a DNA walker was designed for the simultaneous signal amplification of nucleolin (NCL) and thymidine kinase 1 (TK1) in a single cell. NCL and TK1 in single cells were simultaneously detected on a microchip platform with detection limits of 1.0 and 0.8 pM, respectively. The results obtained from 20 liver cancer cells (HepG2) and 20 normal hepatocytes (HL-7702) indicated that NCL and TK1 were overexpressed in liver cancer cells. However, the levels of NCL and TK1 in normal hepatocytes are only about one-tenth to one-sixth of those in hepatic carcinoma. Using different nucleic acid aptamers, the proposed strategy can be applied for the analysis of other single-cell proteins and in the early diagnosis of cancer.

Absolute Quantification of MicroRNAs in a Single Cell with Chemiluminescence Detection Based on Rolling Circle Amplification on a Microchip Platform.

The absolute quantification of miRNAs in a single cell allows to better understand the heterogeneity of cells and the relationship between miRNAs and diseases. However, seldom methods for miRNA quantification in a single cell have been reported because the miRNA content in a single cell is very low. Herein, an ultrasensitive chemiluminescence assay strategy based on rolling circle amplification (RCA) on a microchip platform was proposed for the absolute quantification of miRNAs in a single cell. In this strategy, a ring probe with specificity was designed and synthesized, which could perform RCA for target miRNAs to improve the sensitivity and satisfy the need of absolute quantification of miRNAs in a single cell. The 20 liver cancer cells (HepG2) and 20 normal liver cells (HL-7702) were analyzed using this method; it is found that the miRNA-21 contents varied among cells, and miRNA-21 was overexpressed in HepG2 cells. Compared with traditional methods, the proposed strategy has many advantages such as low cost, simple operation, short analysis time, good specificity, and lower probability of false positives. This method is expected to be one of the powerful tools for the absolute quantification of miRNAs in a single cell.

A New One-Pot Fluorescence Derivatization Strategy for Highly Sensitive MicroRNA Analysis.

MicroRNAs (miRNAs) modulate the expression of over 30 % of mammalian genes during development and apoptosis, and abnormal expression of miRNAs may lead to a range of human pathologies. Therefore, analysis of miRNAs is valuable for disease diagnostics. In this work, a novel one-pot fluorescence derivatization strategy was developed for miRNA analysis. The mechanism of the derivatization reaction was explored by using instrumental methods, including liquid chromatography, fluorescence spectroscopy, and mass spectrometry. Highly fluorescent N6 -ethenoadenine (ϵ-adenine) was formed and detached from the miRNA sequence through the reaction of adenine in nucleic acids with 2-chloroacetaldehyde (CAA) at 100 °C. This is the first experimental evidence that the cooperation of formed ϵ-adenine and water-mediated hydrogen-bond interaction between the proton at the 2'- and the oxyanion at 3'-positions stabilized the oxocarbenium significantly, which makes the depurination and derivatization of miRNA highly effective. Based on this derivatization strategy, a facile and sensitive high-performance liquid chromatography method was developed for quantitative assay of miRNAs. In combination with magnetic solid-phase extraction (MSPE), the HPLC method was shown to be useful for the determination of microRNAs at sub-picomolar level in serum samples.

Mass spectrometric quantification of microRNAs in biological samples based on multistage signal amplification.

This work describes a novel method for quantification of miRNAs based on multistage signal amplification (MSA) and liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). The multistage signal amplification involves hybridization enrichment of miRNA targets with a DNA probe-magnetic bead conjugate, target recycling amplification with a duplex-specific nuclease, and acid hydrolysis of the reporter molecules producing free nucleobases. Nucleobases thus generated are quantified by LC-ESI-MS/MS with specificity and repeatability. Taking miR-21 as the model target, biological samples such as serum and cell cultures were analyzed by using the present protocol. The analytical results indicate that facile and cost-effective quantifications of miRNA targets can be achieved by using the popular LC-ESI-MS/MS technique, and very importantly, without an isolation of total RNAs from the sample prior to the quantitative assay. The assay for miR-21 detection had a linear calibration curve in the range from 0.2 pM to 0.25 nM with a limit of detection of 60 fM. Analysis of MCF-7 cells treated with toremifene (a potent inhibitor of breast cancer cell growth) revealed that the content of miRNA-21 decreased by ca. 50%, and the decrease was dose-dependent.

A G-quadruplex/hemin DNAzyme-based microchip electrophoresis chemiluminescence assay for highly sensitive detection of biotin in flour.

Quantitative analysis of biotin in biological fluids, foods, and pharmaceutical is important for diagnosis and treatment of biotin-related diseases and health maintenance. In this work, a novel G-quadruplex/hemin DNAzyme-based microchip electrophoresis chemiluminescence (CL) assay method was established for rapid and highly sensitive detection of biotin. This method is based on the specific binding between biotin and streptavidin, the catalytic CL characteristics of G-quadruplex/hemin DNAzyme to the oxidation-reduction reaction of hydrogen peroxide with luminol, and the on-line separation function of microchip electrophoresis. Under the optimal experimental conditions, on-chip biotin analysis was achieved within 1 min. The CL intensity is linearly proportional to the concentration of biotin in the range of 13-630 nM with a detection limit of 6.4 nM. The proposed method has been applied for the detection of biotin in flour, biotin contents in three flour samples are found in the range of 199-223 ng/g with a mean value of 214 ng/g. The recoveries were in the range of 94-103%. With excellent sensitivity and good selectivity, the proposed method could be applied in a wide range of biological fluids, foods, and pharmaceutical analysis.

A sensitive fluorescence assay of organophosphorus pesticides using acetylcholinesterase and copper-catalyzed click chemistry.

Organophosphorus pesticides (OPs) are widely used in agricultural fields, but exhibit high toxicity to human beings. A sensitive fluorescence assay for organophosphorus pesticides was developed using the inhibition of acetylcholinesterase (AChE) activity and the copper-catalyzed click chemical reaction. In the click reaction, two hybridized DNA probes can be ligated with copper ions, inducing a fluorescence quenching during the strand displacement reaction. AChE can hydrolyze acetylthiocholine (ATCh) to form thiocholine (TCh) which contains a thiol group. TCh will react with copper ions, blocking the click reaction and a high fluorescence signal is observed. But in the presence of OPs, the activity of AChE is inhibited, releasing a high concentration of copper ions that catalyze the click chemical reaction and resulting in decreased fluorescence signals. Taking advantage of the copper-mediated signal amplification effect, the sensitivity was improved. This assay has also been applied to detect OPs in river water samples with satisfactory results, which demonstrates that the method has great potential for practical applications in environmental protection and food safety fields.

Capsicum-Derived Biomass Quantum Dots Coupled with Alizarin Red S as an Inner-Filter-Mediated Illuminant Nanosystem for Imaging of Intracellular Calcium Ions.

Calcium ion (Ca2+) plays crucial roles in the signal transduction pathways associated with various physiological and pathological events. Monitoring intracellular Ca2+ is of great significance for cell biology research. Here, we report the use of biomass quantum dots (BQDs) as a fluorescent reporter for imaging of intracellular Ca2+, based on the inner-filter-mediated luminescence which was assisted by a Ca2+ chelator alizarin red S (ARS). BQDs were prepared by hydrothermal heating of capsicum. The absorption of ARS overlaps with the excitation of the BQDs, the fluorescence of BQDs being quenched through the inner-filter effect. But the absorption of Ca-ARS complex red shifts and shows a poor inner-filter effect. Thus, Ca2+ can be detected by the inner-filter-mediated luminescence using the BQDs-ARS nanohybrid system. Using the proposed nanosystem, the imaging of intracellular Ca2+ and real-time monitoring of the Ca2+ level change under histamine stimulation were also achieved. Thus, this nanosystem holds potential applications in other Ca2+-related signal transduction study.

An ultrasensitive microchip electrophoresis assay based on separation-assisted double cycling signal amplification strategy for microRNA detection in cell lysate.

Microchip electrophoresis (MCE) assay is an analysis technique with low consumption and high automation. It is a useful tool in biomedical research and clinical diagnosis. However, the low detection sensitivity limits its application in trace biomarker analysis because of its extremely small sample size. To address the need for high sensitivity in MCE, we have developed an ultrasensitive MCE method based on a separation-assisted double cycling signal amplification strategy for the detection of microRNA (miRNA) in cell lysate. In this method, two short single-stranded DNAs P1 and P2 complement each other to form a duplex DNA probe (P1/P2). In the presence of target miRNA, P2 in the P1/P2 probe can be displaced to form double-stranded miRNA/P1. Then, the degradation of P1 in miRNA/P1 by T7 Exo releases the miRNA, and the released miRNA participates in a displacement reaction with another P1/P2 probe to complete the first cycle. The displaced free P2 hybridizes with the hairpin fluorescence probe (MB) to form the P2/MB duplex, which can also be degraded by T7 Exo to release P2. The released P2 can bind with another MB probe to complete the second cycle. By using MCE-laser-induced fluorescence (LIF) as separation and detection platform and miRNA-141 as model analyte, the proposed MCE assay can detect miRNA-141 at concentrations as low as 8.0 fM, which is the highest sensitivity achieved to date for an MCE assay. This method for detecting trace miRNA holds great potential in biomedical research and clinical diagnosis.

Unique Approach To Develop Carbon Dot-Based Nanohybrid Near-Infrared Ratiometric Fluorescent Sensor for the Detection of Mercury Ions.

The ratiometric fluorescence assay, which can eliminate the external effects, has attracted great attention. In this work, a carbon dot (CD)-based nanohybrid dual-emission system was simply prepared by a unique approach of solvothermal treating corn bract and used as a ratiometric fluorescent sensor for Hg2+ detection. Under a single excitation, the obtained nanohybrid sensor had two emission bands around 470 and 678 nm, which may originate from the intrinsic structure of CDs and chlorophyll-derived porphyrins, respectively. In the presence of Hg2+, the fluorescence at 678 nm could be remarkably quenched, while the fluorescence intensity at 470 nm was only slightly altered. The fluorescence intensity ratio at 470 and 678 nm exhibited a good linear relationship in the Hg2+ concentration range from 0 to 40 µM with a detection limit of about 9.0 nM. It also had a satisfying assay performance in serum and river water samples. The prepared CD-based nanohybrid sensor here may hold the further potential applications in biomedicine study, environmental protection, and food safety.

A highly sensitive capillary electrophoresis immunoassay strategy based on dual-labeled gold nanoparticles enhancing chemiluminescence for the detection of prostate-specific antigen.

An enzyme and antibody dual labeled gold nanoparticles enhancing chemiluminescence strategy was developed for highly sensitive CE immunoassay (IA) of prostate-specific antigen (PSA). In this work, gold nanoparticles were labeled with horseradish peroxidase and antiprostate specific antigen-antibody, and used as the marker (Ab* ). After PSA (antigen, Ag) was added into the system, a noncompetitive immune reaction was happen between Ab* and Ag to form an immune complex (Ag-Ab* ). Subsequently, the obtained Ag-Ab* and unreacted Ab* were separated by CE, and the chemiluminescence intensity of Ag-Ab* was used to estimate PSA concentration. The calibration curve showed a good linearity in the range of 0.25-10 ng/mL. Based on a S/N of 3, the detection limit for PAS was estimated to be 0.092 ng/mL. Proposed CE method was applied for PSA quantification in human serum samples from healthy volunteers and patients with prostate cancer. The obtained results demonstrated that the proposed CE method may serve as an alternative tool for clinical analysis of PSA.

Oligonucleotide-stabilized fluorescent silver nanoclusters for the specific and sensitive detection of biotin.

A novel biotin fluorescent probe based on oligonucleotide-stabilized silver nanoclusters (DNA-AgNCs) was synthesized by employing a biotinylated cytosine-rich sequence as a synthesized template. The fluorescence properties of the DNA-AgNCs are related to the modified position of the DNA. When biotin is linked to the middle thymine base of the DNA sequence, the DNA-AgNCs emit the strongest fluorescence. Moreover, the stability of the DNA-AgNCs was affected by avidin through biotin-avidin binding, quenching the fluorescence of the DNA-AgNCs. In contrast, if free biotin is further introduced into this system, the quenching is apparently weakened by competition, leading to the restoration of fluorescence. This phenomenon can be utilized for the detection of biotin. Under the optimal conditions, the fluorescence recovery is linearly proportional to the concentration of biotin in the range of 10 nM-1.0 µM with a detection limit of 6.0 nM. This DNA-AgNCs probe with excellent fluorescent properties is sensitive and selective for the detection of biotin and has been applied for the determination of biotin in wheat flour.

A sensitive strategy for the fluorescence detection of DNA methyltransferase activity based on the graphene oxide platform and T7 exonuclease-assisted cyclic signal amplification.

In this work, a simple fluorescence strategy based on the graphene oxide (GO) platform and T7 exonuclease (T7 Exo)-assisted cyclic signal amplification is developed for the fast and sensitive detection of DNA methyltransferase (MTase) activity and inhibition. In the sensing design, Dam MTase was used as a model analyte. In the presence of Dam MTase, a hairpin probe (HP) was methylated, and then specially recognized and cleaved by Dpn I endonuclease, releasing a ssDNA fragment. The released ssDNA subsequently hybridized with a FAM-labeled signal probe (DP) to form a duplex with a blunt 5'-terminal of DP and a 4-mer overhang at the 5'-end of the released ssDNA. This would trigger the T7 Exo-assisted cyclic signal amplification by repeating the hybridization and digestion of DP, liberating the fluorophore. The liberated fluorophore could not be adsorbed on the GO surface due to low affinity and the fluorescence signal was retained. In contrast, no enzymatic degradation of the DP occurred in the absence of Dam MTase. Thus the intact DP was then adsorbed on the GO surface, resulting in fluorescence quenching. By combining the efficient digestion ability of T7 Exo and the super fluorescence quenching efficiency of GO, the present strategy exhibits a high signal-to-background ratio, providing a satisfying sensitivity for the Dam MTase activity assay. In addition, this method does not require a specific recognition sequence for enzymatic cyclic amplification and dual labels with fluorophore/quencher pairs, making the design easy and low cost. Furthermore, the proposed method was also applied to assay the inhibition of Dam MTase activity. This approach may offer potential applications in clinical diagnostics, drug screening and some other related biomedical research.

The mechanism of human color vision and potential implanted devices for artificial color vision.

Vision plays a major role in perceiving external stimuli and information in our daily lives. The neural mechanism of color vision is complicated, involving the co-ordinated functions of a variety of cells, such as retinal cells and lateral geniculate nucleus cells, as well as multiple levels of the visual cortex. In this work, we reviewed the history of experimental and theoretical studies on this issue, from the fundamental functions of the individual cells of the visual system to the coding in the transmission of neural signals and sophisticated brain processes at different levels. We discuss various hypotheses, models, and theories related to the color vision mechanism and present some suggestions for developing novel implanted devices that may help restore color vision in visually impaired people or introduce artificial color vision to those who need it.

Doping Engineering To Modulate Surface Plasmon Resonance and Enzyme-like Activities for Enhancing Photoacoustic Imaging-Guided Targeted Cancer Therapy in the Second Near-Infrared Window.

Biological imaging-guided targeted tumor therapy has been a soughtafter goal in the field of cancer diagnosis and treatment. To this end, we proposed a strategy to modulate surface plasmon resonance and endow WO3-x nanoparticles (NPs) with enzyme-like catalytic properties by doping Fe2+ in the structure of the NPs. Doping of the Fe2+ introduced oxygen vacancies into the structure of the NPs, inducing a red shift of the maximum absorption wavelength into the near-infrared II (NIR-II) region and enhancing the photoacoustic (PA) and photothermal properties of the NPs for more effective imaging-guided cancer therapy. Under NIR-II laser irradiation, the Fe-WO3-x NPs produced very strong NIR-II PA and photothermal effects, which significantly enhanced the PA imaging and photothermal treatment effects. On the other hand, Fe2+ in Fe-WO3-x could undergo Fenton reactions with H2O2 in the tumor tissue to generate ·OH for chemodynamic therapy. In addition, Fe-WO3-x can also catalyze the above reactions to produce more reactive oxygen species (ROS) and induce the oxidation of NADH to interfere with intracellular adenosine triphosphate (ATP) synthesis, thereby further improving the efficiency of cancer therapy. Specific imaging of tumor tissue and targeted synergistic therapy was achieved after ligation of a MUC1 aptamer to the surface of the Fe-WO3-x NPs by the complexing of -COOH in MUC1 with tungsten ions on the surface of the NPs. These results demonstrated that Fe-WO3-x NPs could be a promising diagnosis and therapeutic agent for cancer. Such a study opens up new avenues into the rational design of nanodiagnosis and treatment agents for NIR-II PA imaging and cancer therapy.

A Silver-Induced Absorption Red-Shifted Dual-Targeted Nanodiagnosis-Treatment Agent for NIR-II Photoacoustic Imaging-Guided Photothermal and ROS Simultaneously Enhanced Immune Checkpoint Blockade Antitumor Therapy.

Tumor metastasis remains a leading factor in the failure of cancer treatments and patient mortality. To address this, a silver-induced absorption red-shifted core-shell nano-particle is developed, and surface-modified with triphenylphosphonium bromide (TPP) and hyaluronic acid (HA) to obtain a novel nanodiagnosis-treatment agent (Ag@CuS-TPP@HA). This diagnosis-treatment agent can dual-targets cancer cells and mitochondria, and exhibits maximal light absorption at 1064 nm, thereby enhancing nesr-infrared II (NIR-II) photoacoustic (PA) signal and photothermal effects under 1064 nm laser irradiation. Additionally, the silver in Ag@CuS-TPP@HA can catalyze the Fenton-like reactions with H2 O2 in the tumor tissue, yielding reactive oxygen species (ROS). The ROS production, coupled with enhanced photothermal effects, instigates immunogenic cell death (ICD), leading to a substantial release of tumor-associated antigens (TAAs) and damage-associated molecular patterns, which have improved the tumor immune suppression microenvironment and boosting immune checkpoint blockade therapy, thus stimulating a systemic antitumor immune response. Hence, Ag@CuS-TPP@HA, as a cancer diagnostic-treatment agent, not only accomplishes targeted the NIR-II PA imaging of tumor tissue and addresses the challenge of accurate diagnosis of deep cancer tissue in vivo, but it also leverages ROS/photothermal therapy to enhance immune checkpoint blockade, thereby eliminating primary tumors and effectively inhibiting distant tumor growth.

Inhibition of dsDNA-templated copper nanoparticles by pyrophosphate as a label-free fluorescent strategy for alkaline phosphatase assay.

On the basis of the inhibition of double strand DNA (dsDNA)-templated fluorescent copper nanoparticles (CuNPs) by pyrophosphate (PPi), a novel label-free turn-on fluorescent strategy to detect alkaline phosphatase (ALP) under physiological conditions has been developed. This method relies on the strong interaction between PPi and Cu(2+), which would hamper the effective formation of fluorescent CuNPs, leading to low fluorescence intensity. The ALP-catalyzed PPi hydrolysis would disable the complexation between Cu(2+) and PPi, facilitating the formation of fluorescent CuNPs through the reduction by ascorbate in the presence of dsDNA templates. Thus, the fluorescence intensity was recovered, and the fluorescence enhancement was related to the concentration of ALP. This method is cost-effective and convenient without any labels or complicated operations. The present strategy exhibits a high sensitivity and the turn-on mode provides a high selectivity for the ALP assay. Additionally, the inhibition effect of phosphate on the ALP activity was also studied. The proposed method using a PPi substrate may hold a potential application in diagnosis of ALP-related diseases or evaluation of ALP functions in biological systems.

A Hg(2+)-mediated label-free fluorescent sensing strategy based on G-quadruplex formation for selective detection of glutathione and cysteine.

A novel label-free fluorescent strategy for the detection of glutathione (GSH) and cysteine (Cys) is presented. The system consists of two single stranded DNA (ssDNA) with thymine-thymine (T-T) mismatches and used Hg(2+) as a mediator, and N-methyl mesoporphyrin IX (NMM) as the signal reporter. The assay is based on the competitive reaction of Hg(2+) with GSH/Cys and T-T mismatched double stranded DNA (dsDNA). In the absence of the target, two ssDNA containing T-T mismatches react with Hg(2+) to form a T-Hg(2+)-T dsDNA structure in the solution, which hampers the formation of a G-quadruplex structure. However, in the presence of the target, GSH/Cys reacts with Hg(2+) to keep DNA probes in a free single state, resulting in the effective formation of a G-quadruplex structure of the DNA probe (GP). Subsequently, due to the strong interaction between the G-quadruplex structure and NMM, fluorescence was greatly enhanced. This fluorescence strategy does not require any chemical modification, making the assay convenient and cost-effective. This method exhibited a linear relationship between peak fluorescence intensity and concentration of GSH in the range of 10-400 nM with a limit of detection (LOD) of 9.6 nM. A linear range for Cys detection was obtained in the concentration range of 10-500 nM with an LOD of 10 nM. Moreover, the proposed method worked well for the analysis of complex biological samples.