Multicolor Fluorescent Nanoprobe Research Articles

Multicolor fluorescence assay of tetracycline: lanthanide complexed amino clay loaded with copper nanoclusters.

A multicolor fluorescent nanoprobe has been prepared by loading bovine serum albumin-stabilized copper nanoclusters (BSA-Cu NCs) onto amino clay (AC) and grafting Eu3+ with auxiliary ligand citric acid (Cit). Tetracycline (TC) can coordinate with Eu3+ by β-diketone structure and transfer energy to Eu3+ through antenna effect. When the concentration of TC is in the range 0 to 13μM, the blue emission intensity of BSA-CuNCs is basically unchanged, and the red emission of Eu3+ is remarkably enhanced due to the coordination with TC. The emission color gradually changes from blue to red under UV lamp (λ = 365nm). However, when the TC concentration is in the range 13 to 350 μΜ, the internal amino acid residues of BSA sensitize TC, and the emission color gradually changed from red to green. The nanoprobe has rich color, is simple to prepare, portable, and provide a wide detection range. The limit of detection (LOD) is 3.04nM, which could be used for real-time visual analysis of trace TC in actual samples (lake water, milk, honey, and bovine serum albumin). In addition, a visual test paper has been designed and combined with the color scanning APP of a smartphone to complete the qualitative and semi-quantitative test of TC. BSA-Cu NCs were loaded on amino clay and graft Eu3+ to establish ratiometric fluorescent nanoprobe for TC detection. With the increase of TC concentration, the emission color under 365nm UV lamp gradually changed from blue to red and then to green, and the color changed obviously and can be observed by the naked eye. The visual test paper and smartphone application detection sensor were developed to realize rapid, convenient, real-time, and visual detection of TC in actual samples.

Highly selective and multicolor ultrasensitive assay of dipicolinic acid: The integration of terbium(III) and gold nanocluster

A novel multicolor fluorescent nano-probe based on the hybridization of Tb3+ ion with gold nanoclusters (Au NCs) was synthesized to monitor and on-site visual assay of 2,6-pyridinedicarboxylic acid (DPA), a biomarker of bacterial spores. DPA can replace the water molecule in the center of Tb3+ and strongly coordinate with Tb3+ based on the analyte-triggered antenna effect. Simultaneously, the red fluorescence of Au NCs is not influenced after addition of DPA and can be used as steady inside fluorescence reference channel to measure background noise. On this basis, the multicolor fluorescence nano-probe based on Tb3+-doped Au NCs for fast analysis of DPA was fabricated. The linear range of this method is 0 to 12.5 μM and the limit of detection is 3.4 nM, which is well below the quantity of DPA concentration of 60 μM released by the spore transmission dose of anthrax infection. The proposed multicolor fluorescence nano-probe was successfully detecting DPA in actual sample with good sensitivity and specificity. In addition, the visual paper-based nano-probe is designed to detect DPA by using the color scanning application of smart phone. This developed platform possesses abroad application prospects with advantages of effective, convenient carrying, simple operation, good selectivity and repeatability.

Molecular engineering for construction of a novel ONOO−- activated multicolor fluorescent nanoprobe for early diagnosis and assessing treatment of arthritis in vivo

Early diagnosis and assessment of the therapeutic effect of arthritis requires a reliable bioanalytical method to quantitatively and selectively detect the biomarkers peroxynitrite (ONOO−) in inflammatory diseases. Compared with previously reported probes for the specific detection of ONOO−, molecular engineering based on ONOO−-activated multicolor fluorescence nanoprobes will have the advantages of providing multi-channel information and be more suitable for bioimaging in multicomponent complex environments. Herein, for the first time, a fluorescent nanoprobe (CSU-FT) based on fluorescence resonance energy transfer (FRET), which can be activated by ONOO−, was constructed for multicolor fluorescence imaging, diagnosis and treatment of arthritis in inflammatory mice. Specifically, an energy transfer scaffold was constructed by conjugation of a near-infrared (NIR) xanthane fluorophore with a rhodamine B fluorophore and multicolor by ONOO−-modulated, which was then grafted onto sodium chondroitin sulfate (CSNa) to form CSU-FT through self-assembly. This nanoprobe shows a fast response time (<20s), outstanding selectivity and excellent detection limits as low as 11.7 nM. Interestingly, CSU-FT has been successfully used for intracellular multichannel imaging of endogenous ONOO− production as well as for diagnosis and treatment in a mice model of arthritis with impressive results, revealing practical application in physiological and pathological connection between ONOO− and arthritis.

Monitoring the Activation of Caspases-1/3/4 for Describing the Pyroptosis Pathways of Cancer Cells.

Pyroptosis is closely related to inhibiting the occurrence and development of tumors. However, the pyroptosis pathways (PPs) impacted by different stimulants are still unknown. To accurately understand the PP in cancer cells, we designed a multicolor fluorescent nanoprobe (Cas-NP) to monitor the activation of caspases-1/3/4 during pyroptosis. The Cas-NP was prepared by the assembly of three different fluorophores-labeled peptides, specific response to caspases-1/3/4 on Au nanoparticles via the Au-Se bond to in situ monitor caspase-1/3/4 with high selectivity and sensitivity. Moreover, the selenopeptide specific to caspase-4 (Cyanine-5-LEVD-SeH) was synthesized for the first time to overcome the difficulty in commercial synthesis. During the pyroptosis of cancer cells induced by adenosine triphosphate (ATP), only the fluorescence of caspase-1 significantly increases. When the cells are stimulated with lipopolysaccharide (LPS), the fluorescence signals corresponding to caspases-3 and 4 first appear and then the fluorescence of caspase-1 is observed. Furthermore, the inhibitor study indicates that the activated caspase-4 can lead to the activation of caspase-1 after the LPS treatment. We first discovered that caspase-3 is activated during the pyroptosis process stimulated by LPS and further verified the activation sequence of caspases-1/3/4 via visualized fluorescence detection. The study provides an effective tool for understanding complex signaling mechanisms in pyroptosis cells and new ideas to explore useful therapeutic inhibitors based on pyroptosis.

Ultrasensitive and visual detection of tetracycline based on dual-recognition units constructed multicolor fluorescent nano-probe

Ultrasensitive and visual detection of tetracycline antibiotic (TC) residues is of great significance to public health and environmental safety. A novel dual-response ratiometric fluorescent nano-probe (SiQDs-Cit-Eu) has been elaborately tailored for the determination and on-site visual assay of tetracycline, by grafting citric acid and europium (Eu3+) ions onto the surface of silicon quantum dots (SiQDs). The blue-emissive SiQDs (λem = 455 nm) fabricated by a one-step facile method act as both scaffold for coordination with Eu3+ ions and recognition unit for TC owing to the inner filter effect (IFE). The coordinate unsaturated red-fluorescent Eu3+ ions (λem = 617 nm) bond to the surface of SiQDs, serving as the specific recognition element for TC due to the antenna effect. In the presence of TC, the as-synthesized nano-probe exhibits double (λem = 455 and 617 nm) and reverse response signals which are accompanied by a marked color change from blue to purple, and then red, thus achieving ultra-high sensitivity with a detection limit of 7.1 nM and instant visual detection of TC in real samples (milk, honey, lake and river water). Furthermore, smartphone-assisted point-of-care testing platform is also constructed based on nano-probe-immobilized test paper by using the color scanning APP.

A Multicolor Fluorescence Nanoprobe Platform Using Two-Dimensional Metal Organic Framework Nanosheets and Double Stirring Bar Assisted Target Replacement for Multiple Bioanalytical Applications.

Multicolor fluorescence probes can show fluorescence of different colors when detecting different targets, and the excellent feature can create a highly differentiated multicolor sensing platform. However, most of the previously reported multicolor luminescent materials usually suffer from high toxicity and photobleaching, complex preparation procedures, and poor water solubility, which may not be conducive to bioanalytical applications. Two-dimensional metal organic frameworks (2D MOFs), which have large specific surface areas with long-range fluorescence quenching coupled with biomolecular recognition events, have encouraged innovation in biomolecular probing. Here, we propose a 2D-MOF-based multicolor fluorescent aptamer nanoprobe using a double stirring bar assisted target replacement system for enzyme-free signal amplification. It utilizes the interaction between 2D MOFs and DNA molecules to detect multiple antibiotics quickly, sensitively, and selectively. Since 2D MOFs have excellent quenching efficiency for luminescence of fluorescent-dye-labeled single-strand DNA (ssDNA), the background fluorescence can be largely reduced and the signal-to-noise ratio can be improved. When the adsorbed ssDNA formed double helix double-stranded DNA with its complementary ssDNA, its fluorescence can be almost fully recovered. The assay was tested by detecting chloramphenicol (CAP), oxytocin (OTC), and kanamycin (KANA) in biological samples. The developed aptasensor was sufficiently sensitive to detect the antibiotic residues as low as 1.5 pM CAP, 2.4 pM OTC, and 1 pM KANA (S/N = 3). It has been preliminarily used for multicolor imaging of three different antibiotics in fish tissue slices with satisfactory results.

A Ratiometric Fluorescent Nano-Probe for Rapid and Specific Detection of Tetracycline Residues Based on a Dye-Doped Functionalized Nanoscaled Metal–Organic Framework

Tetracycline (TC) residues are harmful to the environment and human body, so it is necessary to develop a highly sensitive probe for rapid detection of tetracycline residues. In the present paper, a novel dye-doped porous metal–organic framework (UiO-66)-based multi-color fluorescent nano-probe was designed for sensitive ratiometric detection of tetracycline (TC). In this probe, dye-molecules doped UiO-66 was used as a fluorescent internal standard, and the externally grafted lanthanide Eu3+ complex was used as response signals. The fluorescence of the Eu3+ complex was selectively enhanced with increasing concentrations of TC, which was accompanied by a visual blue-to-red color switch. The nano-probe had a linear response between 0.1 and 6 μM with a lowest detection limit of 17.9 nM, which was much lower than the maximum residue limits set by the United States Food and Drug Administration (676 nM) and the European Union (225 nM). The applicability of this method in the analysis of actual samples was evaluated by the determination of TC in honey and milk samples, indicating satisfactory recovery and good reproducibility. In addition, a cost-effective paper-based probe for rapid and visual detection of TC was developed by fixing the nano-probe on filter papers. With the help of a smartphone camera to capture the fluorescence color, and chromaticity analysis software, the calculation and analysis of red (R) and blue (B) values can be realized, which has the potential for real-time visual detection of TC.

Real-Time in Situ Visualizing of the Sequential Activation of Caspase Cascade Using a Multicolor Gold-Selenium Bonding Fluorescent Nanoprobe.

The caspase cascade is an ensemble of very important signaling molecules that plays a critical role in cell apoptosis. Real-time monitoring of the upstream and downstream activation relationships of the caspases in the signal pathway is of great significance for understanding the regulatory mechanisms of these signaling molecules in the development of various diseases. Herein, a multicolor fluorescent nanoprobe, GNP-Se-Casp, has been developed based on Au-Se bonding for real-time in situ monitoring caspase- (casp-) 3, 8, and 9 during cell apoptosis. In the real-time fluorescence imaging of apoptotic HeLa cells induced by staurosporine using GNP-Se-Casp, the fluorescence signals corresponding to casp-8 and casp-9 sequentially turn on, followed by the appearance of the fluorescence of casp-3, which visualizes the upstream and downstream relationships of casp-3, -8, and -9. Thus, GNP-Se-Casp is an effective tool for real-time in situ monitoring of caspase cascade activation in the apoptosis process of tumor cells. This design strategy is easily adaptable to in situ detection of other signal molecules, especially those with upstream and downstream activation relationships.

Simultaneous Fluorescence Visualization of Epithelial-Mesenchymal Transition and Apoptosis Processes in Tumor Cells for Evaluating the Impact of Epithelial-Mesenchymal Transition on Drug Efficacy.

The epithelial-mesenchymal transition (EMT) process plays a pivotal role in acquiring invasive and metastatic properties and has been recognized as a crucial driver of epithelial-derived tumor malignancies. It is necessary to determine the role of EMT in promoting or suppressing carcinoma progression through investigating the relationship between EMT and apoptosis. We designed a multicolor fluorescent nanoprobe for simultaneously imaging the epithelial biomarker E-cadherin mRNA, the mesenchymal marker vimentin mRNA, and the apoptotic marker caspase-3. EMT and apoptosis progresses could be visually detected, which were used to study the effect of EMT on apoptosis and further assess the influence of EMT on drug efficacy in different cancer cells. We believe the designed nanoprobe can offer a new strategy for visualizing EMT and apoptosis in tumor cells and will be a promising tool to investigate the efficiency of drugs targeting EMT-related therapies in living cells.

Visualizing Breast Cancer Cell Proliferation and Invasion for Assessing Drug Efficacy with a Fluorescent Nanoprobe.

Hard-to-treat cancers are closely relative to uncontrolled cell proliferation, invasion, and metastasis. Assessing proliferation and invasion properties of tumor cells both in vitro and in vivo is especially important for acquiring reliable information for cancer pathogenesis, drug screening, and therapeutic effect evaluation. Herein, we developed a multicolor fluorescent nanoprobe for simultaneously monitoring breast cancer cells' proliferation marker Ki-67 and invasion marker urokinase plasminogen activator (uPA). After treated with the anticancer drugs tamoxifen and curcumin, the changes in cancer cell proliferation and invasion properties were visually detected and therapeutic effects of corresponding drugs were further assessed in vitro and in vivo. The design of the fluorescent nanoprobe opens up an avenue for investigating unscheduled proliferation, invasion, and metastasis in living cells and in vivo and as such will be a promising tool to screen antitumor drugs and evaluate drug efficiency in an extremely efficient manner.

Simultaneous detection of multiple targets involved in the PI3K/AKT pathway for investigating cellular migration and invasion with a multicolor fluorescent nanoprobe.

We develop a multicolor fluorescent nanoprobe for assessing cellular migration and invasion by simultaneously imaging miRNA-221, PTEN mRNA and MMP-9 involved in the PI3K/AKT pathway which can regulate cellular mobility and invasiveness.