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γ-glutamyltransferase (GGT), an important tumor marker, is highly expressed in tumor tissues, and precise detection of its activity provides a vital indicator for the diagnosis and treatment. In this work, a "lighting-on" probe (TCF-GGT) was elaborated to detect endogenous GGT with high selectivity and sensitivity. Dicyanomethyldifuranyl (TCF-OH) was employed as the fluorescence reporter and short peptide glutathione (GSH) worked as the GGT-active trigger, the introduction of which prevented the initial proton transfer of TCF-OH contributing to a blank sensing background. A bright red fluorescence could be switched on upon GGT catalytic hydrolysis, avoiding the potential interference from background. There displayed an excellent water-solubility, and little organic solvent was required during the exploration, which otherwise avoided the potential damage to enzyme and organism. TCF-GGT has been proved to be workable at cellular and organism level with highly effective imaging and a short metabolic cycle, which is expected to offer an alternative solution or reference to the early diagnosis and treatment of tumor., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Rheumatoid arthritis (RA) is well-known as a kind of autoimmune disease, which brings unbearable pain to the patients by multiple organ complications besides arthritis. To date, RA can be hardly cured, but early diagnosis and standard treatment can relieve symptoms and pain. Therefore, an effective tool to assist the early diagnosis of RA deserves considerable attention. On account of the overexpressed ONOO - during the early stage of RA, a near-infrared (NIR) receptor, Lyso-Cy, is proposed in this work by linker chemistry to expand the conjugated rhodamine framework by cyanine groups. Contributed by the pH-sensitive spiral ring in rhodamine, receptor Lyso-Cy has been found to be workable in lysosomes specifically, which was confirmed by the pH-dependent spectra with a narrow responding region and a well-calculated p K a value of 5.81. We presented an excellent ratiometric sensing protocol for ONOO - in an acidic environment, which was also available for targeting ONOO - in lysosomes selectively. This innovative dual-targeting responsive design is expected to be promising for assisting RA diagnosis at an early stage with respect to the joint inflammatory model established in this work at the organism level.

Alzheimer's disease (AD) has become a global threat to the elderly health with a short survival time after diagnosis. Due to the asymptomatic stage during the early development, patients are usually diagnosed at the middle or late stage. Therefore, an efficient tool for AD early diagnosis deserves considerable attention, which could make a significant contribution to the treatment intervention. A fluorescent probe has been widely applied for detecting and visualizing species of interest in vitro and in vivo , and the proper reaction between the probe and analytes is responsible for the fluorescence change to provide a lighting-on or ratiometric responsive pattern with satisfactory sensing behavior. In this work, we report the first attempt to build up an AND-logic probe P2 for AD accuracy diagnosis taking butyrylcholinesterase (BChE) and reactive oxygen species (ROSs) as dual targets. Upon the co-stimulation by these two factors through enzymatic hydrolysis and redox reaction, the NIR emission could be readily turned on. This AND sensing pattern avoided the false-positive response effectively, and other diseases sharing one biomarker could hardly induce a NIR fluorescence response. The sensing assay has also been confirmed to be feasible in vitro and in vivo with good sensibility and selectivity. It is worth mentioning that the probe structure has been optimized in terms of the linkage length. This study shows that probe P2 with a connecting arm of medium length (one methylene, n = 1) has superior sensing performance, promising to provide a reference for the relative structure design.

MicroRNA-21 is an important biomarker of tumor early prediction and metastasis, and its accurate detection is of great significance for tumor diagnosis and treatment. It will be a meaningful work to combine the detection of RNA with chemotherapy and photothermal therapy on the same composite material. Herein, we designed a multifunctional nanocomposite based on gold nanorods (AuNRs), making use of microRNA-triggered drug release and near-infrared photothermal effect, which has been developed for cancer therapy and microRNA-21detection. Firstly, the AuNRs with photothermal effect were synthesized as carriers for drug delivery. Then the surface of gold nanorods was modified by functional DNA chains to provide an efficient site for doxorubicin (DOX) loading. Finally, folic acid was introduced to achieve the targeted treatment of MCF-7 cells. The microRNA competed with the double-stranded DNA, resulting in the release of DOX and the recovery of fluorescence signal located at 595 nm with an excitation of 488 nm effectively. The nano-biosensor could not only achieve dual-function of diagnosis and treatment of cancer cells, but also accomplish the detection of microRNA in tumor cells. It showed a high selectivity for microRNA-21 determination with a limit of detection (LOD) of 2.1 nM from the linear relationship from 1.0 × 10 -5  M to 5.0 × 10 -7  M. This scheme provides an outstanding strategy for cell imaging, treatment, and detection, which serves as a promising candidate in the field of biomedical research., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Homocysteine (Hcy) is one of the important biomarkers of clinical diagnosis, which is closely related to the occurrence and development of many diseases. Current analysis methods have difficulties in detecting Hcy in cells and living organisms. As a powerful technique, fluorescence methods combined the laser confocal imaging technology can achieve real-time visual tracking in cells and in vivo . Herein, we establish a conjugated copolymer-based fluorescence nanosensor (DPA-PFNP-Cu(II)) using the connected 2,7-dibromofluorene and 4,7-bis (2-bromothiophen-5-yl)-2-1-3-benzothiadiazole as the main chain. The competitive coordination between Hcy and Cu(II) allows the fluorescence of the polymer off to on. Finally, the nanosensor is applied for in situ imaging of Hcy levels in the kidney and liver of diabetic mice and is found that Hcy levels were positively correlated with the degree of diabetes. Notably, the depth of tissue penetration of the nanosensor enables Hcy detection of the liver and kidney through in vivo imaging without damage. Two-photon imaging and in vivo imaging achieve consistent results, which correct each other, improving the accuracy of the test result. The present works provide a new imaging technique for studying the occurrence and development of diabetes and screening of new drugs for treatment at the living level.

The modular nature of metal-organic frameworks (MOFs) permits their tunable structure and function for target application, such as in biomedicine. Herein, a green-emission Zr(IV)-MOF (BUT-88) was constructed from a customized luminescent carbazolyl ligand. BUT-88 represents the first bcu-type MOF with both organic linker and metal node in eight connections and shows medium-sized pores, rich accessible linking sites, and good water stability and biocompatibility. In virtue of these merits, BUT-88 was then fabricated into a MOF-based fluorescent nanoprobe, drDNA-BUT-88. Using it, the live-cell imaging of dual tumor biomarkers was achieved for the first time upon a MOF-based probe, offering enhanced detection precision in early cancer diagnosis. Particularly, the probe showed efficient ratiometric fluorescent sensing toward the cytoplasmic biomarker microRNA-21, further improving the detection accuracy at the cellular level. In this work, the elaborate combination of MOF engineering and the fluorescent detection technique has contributed a facile biosensing platform, unlocking more possibilities of MOF chemistry.

A novel background-free fluorescent sensory receptor, with the potential to enable an in situ sensing strategy with new chromophores generated upon the detection event, was designed for the detection of glutathione by forming a fancy 16-ring fluorescent product with one/two-photon excited fluorescence.

The present study shows that a dual-signal nanoprobe consisting of DNAzyme-functionalized porous carbon nanospheres (PCNs) responds to microRNA-21 and zinc ion (Zn 2+ ). The fluorescent probe undergoes an increase in the fluorescence intensity of fluorescein isothiocyanate (FITC) (with excitation/emission wavelengths at 488/517 nm) and the fluorescence intensity of cyanine-5 (Cy5) (with excitation/emission wavelengths at 633/670 nm) in the presence of microRNA-21 and Zn 2+ . The recognition between microRNA-21 and its complementary strand in the PCNs induces the separation of Zn 2+ -specific DNAzyme from PCNs, thus resulting in the increase of green fluorescence, and the exogenous Zn 2+ triggers the rupture of cleavage strand of DNAzyme and recovery of red fluorescence. This nanoprobe allows us to acquire in vitro the determination of microRNA-21 in the range of 2-300 nM with a detection limit of 0.57 nM and the determination of Zn 2+ in the range 2-100 nM with a detection limit of 0.43 nM, and in situ simultaneous imaging in MCF-7 breast cancer cells. Therefore, this strategy permits to obtain the expression levels of different biomarkers in living cells, providing a useful tool for diagnosis of cancers and understanding their biological process. Graphical abstract Schematic representation of the DNAzyme-functionalized porous carbon nanospheres for the imaging analysis of microRNA-21 and Zn2+ in living cells.

Acetylcholinesterase (AChE) is an important enzyme associated with many nervous diseases, demonstrating the great need for smarter sensing platform with improved sensitivity, selectivity and simplified operation. A "turn on" fluorometric assay is described herein for AChE activity detection, according to the specific enzyme catalyzed reaction of acetylcholine (ATCh) by AChE, which generates thiocholine (TCh) as the product. The well-designed fluorescent probe HBTP possesses ESIPT (Excited State Intramolecular Proton Transfer) nature, leading to a larger Stokes shift, which could be quenched upon coordination with Cu 2+ . The fluorescence-silent HBTP-Cu 2+ complex could be broken by TCh generated from reaction of ATCh with AChE, giving rise to HBTP release which originates from competitive coordination of TCh with Cu 2+ . This complex probe HBTP-Cu 2+ offers a limit detection as low as 0.02 mU mL -1 , which is lower than most reported literatures. Furthermore, both HBTP-Cu 2+ and HBTP show little toxicity to live cells and is available in visualizing cellular AChE activity., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Accurate detection of human serum albumin (HSA) in biological samples is quite meaningful for early disease diagnosis and treatment. Herein, a novel fluorescent probe 1-ethyl-4-[2-[4-(diethylamino)-2-hydroxyphenyl]ethenyl]]-pyridinium salt (DEHP) was developed for HSA determination. The inherent fluorescence of DEHP is essentially negligible at physiological conditions assigned to the well-developed twisted intramolecular charge transfer (TICT) protocol. An intriguing fluorescence light up is triggered as the addition of HSA, on account of the inhibited TICT procedure when DEHP enters the hydrophobic cavity of protein HSA. This combination leads to a turn on fluorescent response for HSA with a detection limit of 4.8 nM. After an overall investigation, it has been proved that the strong binding between DEHP and HSA is specific-site-related. In additional, the probe implies a great potential to assist clinical diagnosis due to the usage in actual serum detection. Cell imaging also shows that the probe is expected to monitor HSA production process at cell level., (Copyright © 2019. Published by Elsevier B.V.)

Melanoma is an aggressive malignant tumor that undergoes rapid growth and metastasis in a short time; tyrosinase (TYR) is an important biomarker for melanoma diagnosis as it is over-expressed in melanoma cells. Therefore, the detection of TYR activity is of great significance. Although several fluorescent probes have been reported for the determination of tyrosinase activity in vitro and in vivo, only few of them possess a ratiometric pattern to provide built-in self-calibration for signal correction. Herein, a novel cyanine-based fluorescent probe (Cy-tyr) for the ratiometric fluorescent detection of TYR activity was developed by virtue of the aggregation protocol as a new advancement in this field of melanoma detection. When TYR was applied to the probe Cy-tyr, the phenolic hydroxyl group was oxidized to achieve o-benzoquinone in the presence of oxygen, and the resulting cyanine products exhibited a strong tendency to undergo H-aggregation. Accordingly, the maximum absorption of this probe shifted from 630 nm to 516 nm, and a red fluorescence appeared at 560 nm accompanied by the diminishing of the near infrared (NIR) emission at 760 nm. The proposed ratiometric fluorescent probe showed a good signal-to-noise ratio, leading to high sensitivity for TYR activity with the LOD of 0.02 U mL -1 . Moreover, the probe was successfully applied to image endogenous TYR activity in the B16 cells and showed a complete lack of noise in other cells with lower TYR expressions.

A chromophore is essential to a classic molecular probe, but its intrinsic optical properties can trigger a spectral background, which is considered to produce a negative influence on detection sensitivity. Hence, developing spectral sensing assays with chromophores generated in situ by probes and analytes could realize detection. Herein, we elaborated a novel proposal based on the in situ condensation reaction between 4-diethylaminobenzaldehyde (DEASA) and hydrazine readily promoted by CTAB micelles, resulting in new chromophores with intramolecular hydrogen bonds generated in situ acting as indicators for hydrazine. A background-free detection pattern was, therefore, formulated, along with a relatively low detection limit of 0.42 nM. This avoided tedious preparation and the use of organic solvents during the experiments. In addition, extensive studies, including actual water/serum samples, living cells, and even mouse imaging, revealed that the DEASA probe and the well-designed in situ sensing scheme exhibited good performance regarding future applications in terms of sensitivity, anti-interference capability, and imaging stability.

The effective discrimination of dopamine (DA) analogues is an enduring challenge because of their very tiny structural differences, and thus a separation technique is generally required during the conventional analysis. In this study, a hyperbranched polyethyleneimine (hPEI)-based fluorescent sensor array has been constructed for the separation-free and effective differentiation of four DA analogues. The discrimination includes two steps: firstly, the formation of fluorescent polymer nanoparticles (FPNs) with diverse emission profiles via hPEI-mediated self-polymerization reaction of DA analogues and secondly, the linear discriminant analysis of fluorescence patterns of the formed FPNs for the differentiation of DA analogues. The hPEI-assisted self-polymerization reaction of DA analogues and substitution group mediated optical properties of the resulted FPNs enable an excellent discrimination of four DA analogues at a concentration of 1.0 μM when linear discriminant analysis and hierarchical cluster analysis are smartly combined. Additionally, binary, tertiary and even quaternary mixtures of analogues can also be well distinguished with the proposed sensor array. The practicability of this established sensor array is validated by a high accuracy (100%) evaluation of 88 blind samples containing a single analogue or a mixture of two, three or four analogues.

Chronic hypoxic stress disrupts the intracellular redox homeostasis, leads to a series of physiological dysfunction, and finally results in many diseases including cancer and inflammatory and cardiovascular diseases. The intracellular redox status is related to the homeostasis between reactive oxygen species (ROS) and cellular antioxidant species. Superoxide anion (O 2 •- ) is considered to be a precursor of ROS. As a member of reactive sulfur species, hydrogen polysulfides (H 2 S n ) are a class of antioxidants in cells, which act as an important regulator for the intracellular redox state. Therefore, trapping the cross-talk of O 2 •- and H 2 S n is a benefit for further understanding the physiological and pathological effects. Herein, we conceive a fluorescent probe HCy-ONO for sequential detection of O 2 •- and H 2 S n in cells and in mouse models. Based on a tandem reaction, the probe HCy-ONO can be used to detect O 2 •- and H 2 S n in different fluorescence collection windows without spectral overlap interference with limits of detection 90 and 100 nM, respectively. The strategy affords high sensitivity and selectivity for our detection in living cell models under continuous hypoxic and intermittent hypoxic conditions, revealing the reason for ischemia-reperfusion injury. Moreover, the probe can distinguish the inflamed tissue from normal tissue in acute peritonitis mouse model. Finally, our probe is successfully applied for imaging of O 2 •- and H 2 S n in the SH-SY5Y tumor-bearing mouse model, which is helpful to elucidate the physiological and pathological processes. These data demonstrated that different hypoxic status lead to different concentrations between H 2 S n and O 2 •- .

Herein, we have designed a ratiometric fluorescent nanoprobe for adenosine triphosphate sensing and imaging in living cells, based on silica nanoparticles and a DNA-functionalized hybrid hydrogel. This ratiometric detecting method could validly avoid false-positive signals. Due to its controllable size, favorable biocompatibility and biostability, the nanohydrogel exhibited high cellular permeability and fast response in living cells.

Fluorescent probe-based analytical methods for biological species have gained increasing attention for their powerful detection and imaging capabilities. However, it is still occasionally problematic for accurate analyses due to the influence of the probe concentration, intrinsic fluorescence, instrumental factors or environmental conditions, and intense laser irradiation is usually harmful to biological systems. An NIR ratiometric fluorescent probe can eliminate the influence of these factors and lead to a higher sensing performance. We propose the introduction of a reasonable masking group onto the meso oxygen atom of a cyanine framework (Cy-O) to achieve a NIR ratiometric probe by modulating the conjugated polymethine π-electron system of the cyanine dye. Herein, we report a proof-of-the-concept attempt for a highly sensitive and selective mito-specific NIR ratiometric fluorescent sensing strategy for ALP, and the phosphate group was chosen as the functional group for ALP catalyzed hydrolysis. Upon treatment with ALP, the probe Cy-OP exhibits excellent recognition properties such as high selectivity, high sensitivity (with a low detection limit of 0.16 mU mL -1 ), short response time (within 30 min) and a large spectral shift (from 736 to 516 nm in the absorption spectra and 766 to 616 nm in the emission spectra) with a low fluorescence background. Moreover, the ratio of emission (I 616nm /I 766nm ) of the enzymatic residue was favorable for its application in a complex bio-system. Consequently, the well-designed probe was successfully applied for the detection of ALP in the living cells with satisfactory results.

Gold nanoclusters (AuNCs) protected with a bovine serum albumin (BSA) coating are known to emit red fluorescence (peaking at 650 nm) on photoexcitation with ultraviolet light (365 nm). On addition of Cu(II) ions, fluorescence is quenched because Cu(II) complexes certain amino acid units in the BSA chain. Fluorescence is, however, restored if pyrophosphate (PPi) is added because it will chelate Cu(II) and remove it from the BSA coating on the AuNCs. Because PPi is involved in the function of telomerase, the BSA@AuNCs loaded with Cu(II) can act as a fluorescent probe for determination of the activity of telomerase. A fluorescent assay was worked out for telomerase that is highly sensitive and has a wide linear range (10 nU to 10 fM per mL). The fluorescent probe was applied to the determination of telomerase activity in cervix carcinoma cells via imaging. It is shown that tumor cells can be well distinguished from normal cells by monitoring the differences in intracellular telomerase activity. Graphical abstract Gold nanoclusters (AuNCs) protected by bovine serum albumin (BSA) and displaying red photoluminescence were prepared as fluorescent probe for the determination of telomerase activity and used for imaging of cervix carcinoma (HeLa) cells.

On the basis of the extension reaction of a telomerase substrate (TS) primer in the presence of the telomerase, the inherent signal-transduction mechanism of the hairpin fluorescence probe, and the strand-displacement property of polymerase, an amplified fluorescence detection of telomerase activity in the cancer cells was described in this manuscript. A hairpin fluorescence probe was used as not only the fluorescence signal carrier but also a template of the telomere elongation reaction. In the presence of the telomerase, the stems of the hairpin probes were opened and the telomerase activity could be determined with the fluorescence enhancement. The telomerase activity in the HeLa extracts equivalent to 40-1000 cells was detected by this method, with the multiple rounds of isothermal strand replication, which led to strand displacement, and constituted consecutive signal amplification for the novel detection paradigm that allowed measurement of telomerase activity in crude cell extracts equivalent to 4 cultured HeLa cells. Using magnetic beads as both the separation tool and the immobilization matrix of the aptamer of Ramos cells (CRL-1596, B-cell, human Burkitt's lymphoma), the detection of the amount of the Ramos cell with the low concentration of 100 cells mL(-1) confirmed the reliability and practicality of the protocol, which reveal a good prospect of this platform for analysis.