Molecular histopathology of matrix proteins through autofluorescence super-resolution microscopy.

Extracellular matrix diseases like fibrosis are elusive to diagnose early on, to avoid complete loss of organ function or even cancer progression, making early diagnosis crucial. Imaging the matrix densities of proteins like collagen in fixed tissue sections with suitable stains and labels is a standard for diagnosis and staging. However, fine changes in matrix density are difficult to realize by conventional histological staining and microscopy as the matrix fibrils are finer than the resolving capacity of these microscopes. The dyes further blur the outline of the matrix and add a background that bottlenecks high-precision early diagnosis of matrix diseases. Here we demonstrate the multiple signal classification method-MUSICAL-otherwise a computational super-resolution microscopy technique to precisely estimate matrix density in fixed tissue sections using fibril autofluorescence with image stacks acquired on a conventional epifluorescence microscope. We validated the diagnostic and staging performance of the method in extracted collagen fibrils, mouse skin during repair, and pre-cancers in human oral mucosa. The method enables early high-precision label-free diagnosis of matrix-associated fibrotic diseases without needing additional infrastructure or rigorous clinical training.

Fluorescence imaging sheds light on the immune evasion mechanisms of hepatic stellate cells mediated by superoxide anion.

Whether and how the reactive oxygen species generated by hepatic stellate cells (HSCs) promote immune evasion of hepatocellular carcinoma (HCC) remains mysterious. Therefore, investigating the function of superoxide anion (O2•-), the firstly generated reactive oxygen species, during the immune evasion become necessary. In this work, we establish a novel in situ imaging method for visualization of O2•- changes in HSCs based on a new two-photon fluorescence probe TPH. TPH comprises recognition group for O2•- and HSCs targeting peptides. We observe that O2•- in HSCs gradually rose, impairing the infiltration of CD8+ T cells in HCC mice. Further studies reveal that the cyclin-dependent kinase 4 is deactivated by O2•-, and then cause the up-regulation of PD-L1. Our work provides molecular insights into HSC-mediated immune evasion of HCC, which may represent potential targets for HCC immunotherapy.

High-Spatial-Resolution Benchtop X-ray Fluorescence Imaging through Bragg-Diffraction-Based Focusing with Bent Mosaic Graphite Crystals: A Simulation Study.

X-ray fluorescence imaging (XFI) can localize diagnostic or theranostic entities utilizing nanoparticle (NP)-based probes at high resolution in vivo, in vitro, and ex vivo. However, small-animal benchtop XFI systems demonstrating high spatial resolution (variable from sub-millimeter to millimeter range) in vivo are still limited to lighter elements (i.e., atomic number Z≤45). This study investigates the feasibility of focusing hard X-rays from solid-target tubes using ellipsoidal lens systems composed of mosaic graphite crystals with the aim of enabling high-resolution in vivo XFI applications with mid-Z (42≤Z≤64) elements. Monte Carlo simulations are performed to characterize the proposed focusing-optics concept and provide quantitative predictions of the XFI sensitivity, in silico tumor-bearing mice models loaded with palladium (Pd) and barium (Ba) NPs. Based on simulation results, the minimum detectable total mass of PdNPs per scan position is expected to be on the order of a few hundred nanograms under in vivo conform conditions. PdNP masses as low as 150 ng to 50 ng could be detectable with a resolution of 600 µm when imaging abdominal tumor lesions across a range of low-dose (0.8 µGy) to high-dose (8 µGy) exposure scenarios. The proposed focusing-optics concept presents a potential step toward realizing XFI with conventional X-ray tubes for high-resolution applications involving interesting NP formulations.

Potential application of body fluids autofluorescence in the non-invasive diagnosis of endometrial cancer.

BACKGROUND: Endometrial carcinoma (EC) is the most common cancer of the female reproductive tract in developed countries. The prognosis and 5-year survival rates are closely tied to the stage diagnosis. Current routine diagnostic methods of EC are either lacking specificity or are uncomfortable, invasive and painful for the patient. As of now, the gold diagnostic standard is endometrial biopsy. Early and non-invasive diagnosis of EC requires the identification of new biomarkers of disease and a screening test applicable to routine laboratory diagnostics. The application of untargeted metabolomics combined with artificial intelligence and biostatistics tools has the potential to qualitatively and quantitatively represent the metabolome, but its introduction into routine diagnostics is currently unrealistic due to the financial, time and interpretation challenges. Fluorescence spectral analysis of body fluids utilizes autofluorescence of certain metabolites to define the composition of the metabolome under physiological conditions. PURPOSE: This review highlights the potential of fluorescence spectroscopy in the early detection of EC. Data obtained by three-dimensional fluorescence spectroscopy define the quantitative and qualitative composition of the complex fluorescent metabolome and are useful for identifying biochemical metabolic changes associated with endometrial carcinogenesis. Autofluorescence of biological fluids has the prospect of providing new molecular markers of EC. By integrating machine learning and artificial intelligence algorithms in the data analysis of the fluorescent metabolome, this technique has great potential to be implemented in routine laboratory diagnostics.

Real-time identification of life-threatening necrotizing soft-tissue infections using indocyanine green fluorescence imaging.

Significance: Necrotizing soft-tissue infections (NSTIs) are life-threatening infections with a cumulative case fatality rate of 21%. The initial presentation of an NSTI is non-specific, frequently leading to misdiagnosis and delays in care. No current strategies yield an accurate, real-time diagnosis of an NSTI. Aim: A first-in-kind, observational, clinical pilot study tested the hypothesis that measurable fluorescence signal voids occur in NSTI-affected tissues following intravenous administration and imaging of perfusion-based indocyanine green (ICG) fluorescence. This hypothesis is based on the established knowledge that NSTI is associated with local microvascular thrombosis. Approach: Adult patients presenting to the Emergency Department of a tertiary care medical center at high risk for NSTI were prospectively enrolled and imaged with a commercial fluorescence imager. Single-frame fluorescence snapshot and first-pass perfusion kinetic parameters-ingress slope (IS), time-to-peak (TTP) intensity, and maximum fluorescence intensity (IMAX)-were quantified using a dynamic contrast-enhanced fluorescence imaging technique. Clinical variables (comorbidities, blood laboratory values), fluorescence parameters, and fluorescence signal-to-background ratios (SBRs) were compared to final infection diagnosis. Results: Fourteen patients were enrolled and imaged (six NSTI, six cellulitis, one diabetes mellitus-associated gangrene, and one osteomyelitis). Clinical variables demonstrated no statistically significant differences between NSTI and non-NSTI patient groups (p-value≥0.22). All NSTI cases exhibited prominent fluorescence signal voids in affected tissues, including tissue features not visible to the naked eye. All cellulitis cases exhibited a hyperemic response with increased fluorescence and no distinct signal voids. Median lesion-to-background tissue SBRs based on snapshot, IS, TTP, and IMAX parameter maps ranged from 3.2 to 9.1, 2.2 to 33.8, 1.0 to 7.5, and 1.5 to 12.7, respectively, for the NSTI patient group. All fluorescence parameters except TTP demonstrated statistically significant differences between NSTI and cellulitis patient groups (p-value<0.05). Conclusions: Real-time, accurate discrimination of NSTIs compared with non-necrotizing infections may be possible with perfusion-based ICG fluorescence imaging.

Autofluorescence-Guided Total Thyroidectomy in Low-Volume, Nonparathyroid Institutions.

Importance: Hypoparathyroidism following thyroid surgery is a serious complication that occurs frequently when surgery is performed by low-volume thyroid surgeons without experience in parathyroid surgery. Objective: To evaluate the occurrence of hypoparathyroidism following total thyroidectomy after the introduction of autofluorescence in low-volume, nonparathyroid institutions. Design, Setting, and Participants: This prospective, multicenter cohort study, with a follow-up period of up to 1 year, was conducted in Denmark at 2 low-volume nonparathyroid institutions between January 2021 and November 2023. All adult patients referred for total thyroidectomy were assessed for eligibility (n = 90). Only patients with no history of thyroid surgery were considered (n = 89). Patients who only underwent lobectomy (n = 6) or declined to participate (n = 5) were excluded. All included patients completed follow-up. The prospective cohort was compared with a historical cohort of successive patients undergoing primary total thyroidectomy from 2016 to 2020 (before autofluorescence was available). Intervention: Included patients underwent autofluorescence-guided total thyroidectomy. Main outcomes and Measures: Rate of hypoparathyroidism. Immediate hypoparathyroidism was defined as the need for active vitamin D postoperatively, whereas permanent hypoparathyroidism was considered when there still was a need for active vitamin D 1 year after surgery. Results: Seventy-eight patients underwent autofluorescence-guided surgery (mean [SD] age, 55.6 [13.1] years; 67 [86%] female) and were compared with 89 patients in the historical cohort (mean [SD] age, 49.7 [12.8] years; 78 [88%] female). The rate of immediate hypoparathyroidism decreased from 37% (95% CI, 27%-48%) to 19% (95% CI, 11%-30%) after the introduction of autofluorescence (P = .02). Permanent hypoparathyroidism rates decreased from 32% (95% CI, 22%-42%) to 6% (95% CI, 2%-14%) (P < .001), reaching 0% at the end of the study. More parathyroid glands were identified with autofluorescence (75% [95% CI, 70%-80%] vs 61% [95% CI, 56%-66%]) (P < .001) and less parathyroid glands were inadvertently excised (4% [95% CI, 1%-11%] vs 21% [95% CI, 13%-31%]) (P = .001). Conclusions and Relevance: In this cohort study of autofluorescence-guided thyroid surgery in low-volume, nonparathyroid institutions, the use of autofluorescence was associated with a significant decrease in both immediate and permanent hypoparathyroidism. When autofluorescence was used, hypoparathyroidism rates were comparable with those of high-volume surgeons who also perform parathyroid surgery.

Controlled In Situ Self-Assembly of Biotinylated Trans-Cyclooctene Nanoparticles for Orthogonal Dual-Pretargeted Near-Infrared Fluorescence and Magnetic Resonance Imaging.

A pretargeted strategy that decouples targeting vectors from radionuclides has shown promise for nuclear imaging and/or therapy in vivo. However, the current pretargeted approach relies on the use of antibodies or nanoparticles as the targeting vectors, which may be compromised by poor tissue penetration and limited accumulation of targeting vectors in the tumor tissues. Herein, we present an orthogonal dual-pretargeted approach by combining stimuli-triggered in situ self-assembly strategy with fast inverse electron demand Diels-Alder (IEDDA) reaction and strong biotin-streptavidin (SA) interaction for near-infrared fluorescence (NIR FL) and magnetic resonance (MR) imaging of tumors. This approach uses a small-molecule probe (P-Cy-TCO&Bio) containing both biotin and trans-cyclooctene (TCO) as a tumor-targeting vector. P-Cy-TCO&Bio can efficiently penetrate subcutaneous HeLa tumors through biotin-assisted targeted delivery and undergo in situ self-assembly to form biotinylated TCO-bearing nanoparticles (Cy-TCO&Bio NPs) on tumor cell membranes. Cy-TCO&Bio NPs exhibited an "off-on" NIR FL and retained in the tumors, offering a high density of TCO and biotin groups for the concurrent capture of Gd-chelate-labeled tetrazine (Tz-Gd) and IR780-labeled SA (SA-780) via the orthogonal IEDDA reaction and SA-biotin interaction. Moreover, Cy-TCO&Bio NPs offered multiple-valent binding modes toward SA, which additionally regulated the cross-linking of Cy-Gd&Bio NPs into microparticles (Cy-Gd&Bio/SA MPs). This process could significantly (1) increase r1 relaxivity and (2) enhance the accumulation of Tz-Gd and SA-780 in the tumors, resulting in strong NIR FL, bright MR contrast, and an extended time window for the clear and precise imaging of HeLa tumors.

Novel biocompatible N-rich AIE fluorescent probe for live cell imaging and visual onsite detection of uranium.

A sensitive and biocompatible N-rich probe for rapid visual uranium detection was constructed by grafting two trianiline groups to 2,6-bis(aminomethyl)pyridine. Possessing excellent aggregation-induced emission (AIE) property and the advantages to form multidentate chelate with U selectively, the probe has been applied successfully to visualize uranium in complex environmental water samples and living cells, demonstrating outstanding anti-interference ability against large equivalent of different ions over a wide effective pH range. A large linear range (1.0 × 10-7-9.0 × 10-7 mol/L) and low detection limit (72.6 nmol/L, 17.28 ppb) were achieved for the visual determination of uranium. The recognition mechanism, photophysical properties, analytical performance and cytotoxicity were systematically investigated, demonstrating high potential for fast risk assessment of uranium pollution in field and in vivo.

Optical imaging (HandScan) can identify ultrasound remission in rheumatoid arthritis.

BACKGROUND: Identifying remission is of high importance in rheumatoid arthritis (RA) because remission is associated with less structural progression. We investigated the efficacy of a new optical imaging device, HandScan, to identify RA remission, as defined by ultrasound (US). METHODS: 61 RA patients were included. Disease activity was evaluated by clinical assessment and US, using gray-scale (GS) and Power Doppler (PD). HandScan determined unitary optical spectral transmission (OST) values for wrists, metacarpophalangeal and proximal interphalangeal joints. At the patient level, three composite HandScan (HS) scores were calculated: total HS score; disease activity score OST (DAS-OST) and DAS-OST without patient global assessment (PtGA). Using ROC curves, we determined HS cut-offs to identify US-defined remission. RESULTS: At the joint level, unitary OST values significantly correlated with GS synovitis [odds ratio (OR) 2.43, p < 0.0001] and PD positivity (OR 3.72, p = 0.0002 ). At the patient level, total HS score and DAS-OST were significantly associated with all gray-scale US (GSUS) and power doppler US (PDUS) parameters evaluated (synovitis number and grade, synovial thickness, PD grade) (p < 0.05). The cut-off to identify US-defined remission at the joint level was of 0.92, giving an 81% sensitivity and a 96% positive predictive value (PPV). At the patient level, ROC-curves failed to identify a robust cut-off for the total HS score, but did identify a cut-off (3.68) for DAS-OST to identify US-defined remission, but with lower sensitivity (75%), specificity (56%) and PPV (67%). CONCLUSIONS: HandScan is a non-invasive optical imaging technique providing OST values that correlate with GSUS and PDUS parameters. In addition, HandScan is able to reliably identify US-defined remission in RA at the joint level, with a good sensitivity and high PPV. At the patient level, HandScan DAS-OST can also determine US remission (while total HS score failed to do so), but with lower performance.

A mitochondrion-targeted cyanine agent for NIR-II fluorescence-guided surgery combined with intraoperative photothermal therapy to reduce prostate cancer recurrence.

Poorly identified tumor boundaries and nontargeted therapies lead to the high recurrence rates and poor quality of life of prostate cancer patients. Near-infrared-II (NIR-II) fluorescence imaging provides certain advantages, including high resolution and the sensitive detection of tumor boundaries. Herein, a cyanine agent (CY7-4) with significantly greater tumor affinity and blood circulation time than indocyanine green was screened. By binding albumin, the absorbance of CY7-4 in an aqueous solution showed no effects from aggregation, with a peak absorbance at 830 nm and a strong fluorescence emission tail beyond 1000 nm. Due to its extended circulation time (half-life of 2.5 h) and high affinity for tumor cells, this fluorophore was used for primary and metastatic tumor diagnosis and continuous monitoring. Moreover, a high tumor signal-to-noise ratio (up to ~ 10) and excellent preferential mitochondrial accumulation ensured the efficacy of this molecule for photothermal therapy. Therefore, we integrated NIR-II fluorescence-guided surgery and intraoperative photothermal therapy to overcome the shortcomings of a single treatment modality. A significant reduction in recurrence and an improved survival rate were observed, indicating that the concept of intraoperative combination therapy has potential for the precise clinical treatment of prostate cancer.

A novel ESIPT fluorescent probe for early detection and assessment of ferroptosis-mediated acute kidney injury via peroxynitrite fluctuation.

BACKGROUND: Acute kidney injury (AKI) poses a severe risk to public health, mostly manifested by damage and death of renal tubular epithelial cells. However, routine blood examination, a conventional approach for clinical detection of AKI, is not available for identifying early-stage AKI. Plenty of reported methods were lack of early biomarkers and real time evaluation tools, which resulted in a vital challenge for early diagnosis of AKI. Therefore, developing novel probes for early detection and assessment of AKI is exceedingly crucial. RESULTS: Based on ESIPT mechanism, a new fluorescent probe (MEO-NO) with 2-(2'-hydroxyphenyl) benzothiazole (HBT) derivatives as fluorophore has been synthesized for dynamic imaging peroxynitrite (ONOO-) levels in ferroptosis-mediated AKI. Upon the addition of ONOO-, MEO-NO exhibited obvious fluorescence changes, a significant Stokes shift (130 nm) and rapid response (approximately 45 s), and featured exceptional sensitivity (LOD = 7.28 nM) as well as high selectivity from the competitive species at physiological pH. In addition, MEO-NO was conducive to the biological depth imaging ONOO- in cells, zebrafish, and mice. Importantly, MEO-NO could monitor ONOO- levels during sorafenib-induced ferroptosis and CP-induced AKI. With the assistance of MEO-NO, we successfully visualized and tracked ONOO- variations for early detection and assessment of ferroptosis-mediated AKI in cells, zebrafish and mice models. SIGNIFICANCE AND NOVELTY: Benefiting from the superior performance of MEO-NO, experimental results further demonstrated that the levels of ONOO- was overexpressed during ferroptosis-mediated AKI in cells, zebrafish, and mice models. The developed novel probe MEO-NO provided a strong visualization tool for imagining ONOO-, which might be a potential method for the prevention, diagnosis, and treatment of ferroptosis-mediated AKI.

Hydrazo Pyrazole-Pyridone Fluorescent tag for NLO, Live cell imaging, LFPs visualization, Photophysical probing, and Electrochemical sensor for Dopamine detection.

The present communication reports on the synthesis of a novel methyl-pyridone azo fluorescent tag (MPAFT) were proven through 1H (NMR), FT-IR, UV-vis, and high-resolution mass spectrometry. The quantum chemical parameters of MPAFT were evaluated using density functional theory (DFT) analysis. It was further investigated for its latent fingerprint (LFPs) in various surfaces and anticounterfeiting applications. By exposing Level I-Level III, ridge features to UV light with a wavelength of 365 nm, a bioimaging investigation has also demonstrated the potential of MPAFT's emission behaviour. The cyclic voltammetry (CV) and linear sweep voltammetry (LSV) at MPAFT/MGCE (modified glassy carbon electrode) were used to explore the electrochemical sensitivity and reliable detection of dopamine (DA) in neutral PBS (pH 7) electrolyte solution, and the results show good sensitivity and detection. The lower detection limit for LSV was 0.81 µM under optimum conditions.

A V-shaped bis-coumarin based fluorescence probe for F- detection in tea infusions and potable water and bioimaging applications in living systems.

Fluorine (F) is a pivotal element in the formation of human dental and skeletal tissues, and the consumption of water and tea constitutes a significant source of fluoride intake. However, prolonged ingestion of water and tea with excessive fluoride content can lead to fluorosis, which poses a serious health hazard. In this manuscript, a novel turn-on fluorescent probe DCF synthesized by bis-coumarin and tert-butyldiphenylsilane (TBDPS) was introduced for detecting F- in potable water and tea infusions. By leveraging the unique chemical affinity between fluoride and silicon, F- triggers the silicon-oxygen bond cleavage in DCF, culminating in a conspicuous emission of yellow fluorescence. Validated through a succession of optical tests, this probe exhibits remarkable advantages in terms of superior selectivity, a low detection limit, a large Stokes shift, and robust interference resistance when detecting inorganic fluoride. Moreover, it can serve as portable test strips for on-site real-time identification and quantitative analysis of F-. Furthermore, the application of DCF for in-situ monitoring and imaging of F- in zebrafish and soybean root tissues proved its significant value for F- detection in both animal and plant systems. This probe potentially functions as an efficient instrument for delving into the toxic mechanisms of fluoride in physiological processes.

Acid-Triggered Degradation of Three-In-One Ag2S Quantum Dots for In Situ Ratiometric NIR-II Fluorescence Imaging-Guided Ion/Gas Combination Therapy.

Smart theranostic nanoprobes with the integration of multiple therapeutic modalities are preferred for precise diagnosis and efficient therapy of tumors. However, it remains a big challenge to arrange the imaging and two or more kinds of therapeutic agents without weakening the intended performances. In addition, most existing fluorescence (FL) imaging agents suffer from low spatiotemporal resolution due to the short emission wavelength (<900 nm). Here, novel three-in-one Ag2S quantum dot (QD)-based smart theranostic nanoprobes were proposed for in situ ratiometric NIR-II FL imaging-guided ion/gas combination therapy of tumors. Under the acidic tumor microenvironment, three-in-one Ag2S QDs underwent destructive degradation, generating toxic Ag+ and H2S. Meanwhile, their FL emission at 1270 nm was weakened. Upon introduction of a downconversion nanoparticle (DCNP) as the delivery carrier and NIR-II FL reference signal unit, the formed Ag2S QD-based theranostic nanoprobes could achieve precise diagnosis of tumors through ratiometric NIR-II FL signals. Also, the generated Ag+ and H2S enabled specific ion/gas combination therapy toward tumors. By combining the imaging and therapeutic functions, three-in-one Ag2S QDs may open a simple yet reliable avenue to design theranostic nanoprobes.

A thiazole-based colorimetric and photoluminescent chemosensors for As3+ ions detection: Density functional theory, test strips, real samples, and bioimaging applications.

A Schiff-base Ethyl (E)-2-(3-((2-carbamothioylhydrazono)methyl)-4-hydroxyphenyl)-4-methylthiazole-5-carboxylate (TZTS) dual functional colorimetric and photoluminescent chemosensor which includes thiazole and thiosemicarbazide has been synthesized to detect arsenic (As3+) ions selectively in DMSO: H2O (7:3, v/v) solvent system. The molecular structure of the probe was characterized via FT-IR, 1H, and 13C NMR & HRMS analysis. Interestingly, the probe exhibits a remarkable and specific colorimetric and photoluminescence response to As3+ ions when exposed to various metal cations. The absorption spectral changes of TZTS were observed upon the addition of As3+ ions, with a naked eye detectable color change from colorless to yellow color. Additionally, the chemosensor (TZTS) exhibited a new absorption band at 412 nm and emission enhancements in photoluminescence at 528 nm after adding As3+ ions. The limit of detection (LOD) for As3+ ions was calculated to be 16.5 and 7.19 × 10-9 M by the UV-visible and photoluminescent titration methods, respectively. The underlying mechanism and experimental observations have been comprehensively elucidated through techniques such as Job's plot, Benesi-Hildebrand studies, and density functional theory (DFT) calculations. For practical application, the efficient determination of As3+ ions were accomplished using a spike and recovery approach applied to real water samples. In addition, the developed probe was successfully employed in test strip applications, allowing for the naked-eye detection of arsenic ions. Moreover, fluorescence imaging experiments of As3+ ions in the breast cancer cell line (MCF-7) demonstrated their practical applications in biological systems. Consequently, these findings highlight the significant potential of the TZTS sensor for detecting As3+ ions in environmental analysis systems.

Exogenous and Endogenous Dual-Activated Nanoladder for Precise Imaging of Mitochondrial Ferroptosis-Related Inhibition miRNA with Tumor Cell Specificity.

Developing precise tumor cell-specific mitochondrial ferroptosis-related inhibition miRNA imaging methods holds enormous potential for anticancer drug screening and cancer treatment. Nevertheless, traditional amplification methods still tolerated the limited tumor specificity because of the "off-tumor" signal leakage resulting from their "always-active" sensing mode. To overcome this limitation, we herein developed a dual (exogenous 808 nm NIR light and endogenous APE1) activated nanoladder for precise imaging of mitochondrial ferroptosis-related miRNA with tumor cell specificity and improved imaging resolution. Exogenous NIR light-activation can regulate the ferroptosis-related inhibition miRNA imaging signals within mitochondria, and endogenous enzyme-activation can confine signals to tumor cells. Based on this dual activation design, off-tumor signals were greatly reduced and tumor-to-background contrast was enhanced with an improved tumor/normal discrimination ratio, realizing tumor cell-specific precise imaging of mitochondrial ferroptosis-related inhibition miRNA.

A Red-Emission Fluorescent Probe with Large Stokes Shift for Detection of Viscosity in Living Cells and Tumor-Bearing Mice.

Abnormal viscosity is closely related to the occurrence of many diseases, such as cancer. Therefore, real-time detection of changes in viscosity in living cells is of great importance. Fluorescent molecular rotors play a critical role in detecting changes in cellular viscosity. Developing red emission viscosity probes with large Stokes shifts and high sensitivity and specificity remains an urgent and important topic. Herein, a novel viscosity-sensitive fluorescent probe (TCF-VIS1) with a large stokes shift and red emission was prepared based on the 2-dicyanomethylene-3-cyano-4,5,5-trimethyl-2,5-dihydrofuran (TCF) skeleton. Due to intramolecular rotation, the probe itself does not fluorescence at low viscosity. With the increase in viscosity, the rotation of TCF-VIS1 is limited, and its fluorescence is obviously enhanced. The probe has the advantages of simple preparation, large Stokes shift, good sensitivity and selectivity, and low cytotoxicity, which make it successfully used for viscosity detection in living cells. Moreover, TCF-VIS1 showed its potential for cancer diagnosis at the cell level and in tumor-bearing mice by detecting viscosity. Therefore, the probe is expected to enrich strategies for the detection of viscosity in biological systems and offer a potential tool for cancer diagnosis.