Rapid detection of hypobromous acid by a tetraphenylethylene-based turn-on fluorescent AIE probe and its applications.

BACKGROUND: Similar to hypochlorous acid (HClO), hypobromous acid (HBrO) is one of the most notable reactive oxygen species (ROS). Overexpression of HBrO is linked to various diseases causing organ and tissue loss. Due to HBrO's role in the oxidation of micropollutants, real-time monitoring of HBrO in water-based systems is essential. Tetraphenylethylene (TPE)-based organic aggregation-induced emission luminophores (AIEgens) are an emerging category of fluorescent probe materials that have attracted considerable attentions. However, AIE probes are rarely applied to detect HBrO. Developing faster, more precise, and more sensitive AIE probes is thus crucial for detecting biological and environmental HBrO. RESULTS: A small molecule fluorescent probe 4-(1,2,2-triphenylvinyl)benzamidoxime (SWJT-21) was synthesized for the sensitive and selective detection of hypobromous acid (HBrO) based on aggregation-induced emission (AIE). The amidoxime unit of SWJT-21 would undergo an oxidation reaction with HBrO, leading to a structure differentiation between the probe and the product, and therefore the turn-on fluorescence by the AIE effect. The probe could recognize hypobromous acid rapidly (less than 3 s) in high aqueous phase (99 % water) with a turn-on fluorescence response. It was determined that the limit of detection for HBrO was 5.47 nM. Moreover, SWJT-21 demonstrates potential as a test strip for the detection of HBrO. SWJT-21 was also successfully used for the monitoring of HBrO in water samples and for the detection of endogenous/exogenous HBrO in living cells and zebrafish. SIGNIFICANCE: A special AIE fluorescence turn-on probe SWJT-21 based on tetraphenylethylene was designed for detecting HBrO in the environmental and biological systems. This probe has an extremely low detection limit of 5.47 nM and is able to detect HBrO in 99 % aqueous phase in less than 3 s.

A novel fluorescent indicator-based fluorene derivative for rapid and visual trace water sensing.

Fluorene nucleus derivatives show great potential for building outstanding fluorescence probes. In this paper, a novel fluorescent probe was developed by reacting with fluorene core with azacyclobutane, which exhibits typical solvation chromogenic effect in solvent. The fluorescence of the probe quenched in highly polar solvent. Based on this phenomenon, a novel fluorescence system for trace water was constructed. The response of this probe was fast (30 s) and sensitive for the detection of trace water in organic solvents, and the detection limit of water content in DMSO reached 0.13%. In addition, the probe can also be made as a test strip combined with homemade portable device and a smartphone for rapid detection of trace water. The luminescence mechanism of the probe is theoretically calculated based on time-contained density functional theory (TDDFT). To showcase its practicality, it has been applied for the detection of trace water in honey and alcohol by dipstick. This method provides a new idea for designing efficient fluorescent probes based on dipstick and mobile phone rapid detection.

Coumarin-Synthetic Methodologies, Pharmacology, and Application as Natural Fluorophore.

Coumarins are secondary metabolites made up of benzene and α-pyrone rings fused together that can potentially treat various ailments, including cancer, metabolic, and degenerative disorders. Coumarins are a diverse category of both naturally occurring as well as synthesized compounds with numerous biological and therapeutic properties. Coumarins as fluorophores play a key role in fluorescent labeling of biomolecules, metal ion detection, microenvironment polarity detection, and pH detection. This review provides a detailed insight into the characteristics of coumarins as well as their biosynthesis in plants and metabolic pathways. Various synthetic strategies for coumarin core involving both conventional and green methods have been discussed comparing advantages and disadvantages of each method. Conventional methods discussed are Pechmann, Knoevenagel, Perkin, Wittig, Kostanecki, Buchwald-Hartwig, and metal-induced coupling reactions such as Heck and Suzuki, as well as green approaches involving microwave or ultrasound energy. Various pharmacological applications of coumarin derivatives are discussed in detail. The structural features and conditions responsible for influencing the fluorescence of coumarin core are also elaborated.

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.

Efficient turn-on fluorescent probe cooperated by cascade response for disclosing the fluctuation of cysteine in cells.

BACKGROUND: The research on cysteine (Cys) determination is deemed as a hot topic, since it has been reported to be connected with various physiological processes and disease prediction. However, existing Cys-responding probes may expose some defects such as long reaction time, disappointing photostability, and suboptimal sensitivity. Under such a circumstance, our team has proposed an efficient fluorescent probe with novel sensing mechanism to perfectly cope with the above-mentioned drawbacks. RESULTS: A novel cascade reaction-based probe 9-(2,2-dicyanovinyl)-2,3,6,7-tetrahydro-1H,5H-pyrido[3,2,1-ij]quinolin-8-yl acrylate (DPQA) has been synthesized for the first time. Undergoing addition-cleavage and cyclization-rearrangement processes, DPQA reacts with Cys to generate an iminocoumarin product with relucent green fluorescence, namely 11-imino-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1-ij]quinoline-10-carbonitrile (IMC-J), and the relative fluorescence quantum yield (Φf) soars from 0.007 to 0.793. Utilizing such a mechanism, DPQA shows a superb turn-on signal (172-fold), low detection limit (4.1 nM), and wide detection range (5-6000 nM) toward Cys detection. Encouraged by the admirable sensing performance of DPQA, bioimaging of endogenous Cys has been attempted in HeLa cells with satisfactory results. Moreover, cell model of H2O2-induced oxidative stress has been established and the Cys fluctuation during this process has been inspected, elucidating how living cells confront with the eruption of reactive oxygen species (ROS) storm. SIGNIFICANCE: The probe DPQA with such an intriguing cascade responding process for Cys detection has been endowed with many merits, such as fast reaction and superior sensitivity, conducive to improving responsiveness and rendering it more suitable for further applications. Thereby, we expect that the DPQA would be an efficient tool for detecting Cys fluctuation in living cells of different physiological processes.

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.

Two Fluorescent Probes for Recognition of Acetylcholinesterase: Design, Synthesis, and Comparative Evaluation.

In this study, two "on-off" probes (BF2-cur-Ben and BF2-cur-But) recognizing acetylcholinesterase (AChE) were designed and synthesized. The obtained probes can achieve recognition of AChE with good selectivity and pH-independence with a linear range of 0.5~7 U/mL and 0.5~25 U/mL respectively. BF2-cur-Ben has a lower limit of detection (LOD) (0.031 U/mL), higher enzyme affinity (Km = 16 ± 1.6 µM), and higher inhibitor sensitivity. A responsive mechanism of the probes for AChE was proposed based on HPLC and mass spectra (MS) experiments, as well as calculations. In molecular simulation, BF2-cur-Ben forms more hydrogen bonds (seven, while BF2-cur-But has only four) and thus has a more stable enzyme affinity, which is mirrored by the results of the comparison of Km values. These two probes could enable recognition of intracellular AChE and probe BF2-cur-Ben has superior cell membrane penetration due to its higher log p value. These probes can monitor the overexpression of AChE during apoptosis of lung cancer cells. The ability of BF2-cur-Ben to monitor AChE in vivo was confirmed by a zebrafish experiment.

Real-time monitoring of abnormal mitochondrial viscosity in glioblastoma with a novel mitochondria-targeting fluorescent probe.

Glioblastoma is the most fatal and insidious malignancy, due to the existence of the blood-brain barrier (BBB) and the high invasiveness of tumor cells. Abnormal mitochondrial viscosity has been identified as a key feature of malignancies. Therefore, this study reports on a novel fluorescent probe for mitochondrial viscosity, called ZVGQ, which is based on the twisted intramolecular charge transfer (TICT) effect. The probe uses 3-dicyanomethyl-1,5,5-trimethylcyclohexene as an electron donor moiety and molecular rotor, and triphenylphosphine (TPP) cation as an electron acceptor and mitochondrial targeting group. ZVGQ is highly selective, pH and time stable, and exhibits rapid viscosity responsiveness. In vitro experiments showed that ZVGQ could rapidly recognize to detect the changes in mitochondrial viscosity induced by nystatin and rotenone in U87MG cells and enable long-term imaging for up to 12 h in live U87MG cells. Additionally, in vitro 3D tumor spheres and in vivo orthotopic tumor-bearing models demonstrated that the probe ZVGQ exhibited exceptional tissue penetration depth and the ability to penetrate the BBB. The probe ZVGQ not only successfully visualizes abnormal mitochondrial viscosity changes, but also provides a practical and feasible tool for real-time imaging and clinical diagnosis of glioblastoma.

Construction of a novel near-infrared fluorescent Nile blue@MOF nanoprobe for imaging mitochondrial ATP in living cells.

A near-infrared fluorescent nanoprobe consisting of Nile blue-capped ZIF-90 is first proposed for real-time imaging of mitochondrial ATP. Owing to the strong binding of ATP with Zn2+, the structure of the probe is disrupted, leading to the release of fluorescent NB.

Exploring mitochondrial targeting: an innovative fluorescent probe reveals Nernstian potential and partitioning combination.

This study introduces a paradigm-shifting approach to optimize mitochondrial targeting. Employing a new fluorescent probe strategy, we unravel a combined influence of both Nernst potential (Ψ) and partitioning (P) contributions. Through the synthesis of new benz[e]indolinium-derived probes, our findings redefine the landscape of mitochondrial localization by optimizing the efficacy of mitochondrial probe retention in primary cortical neurons undergoing normoxia and oxygen-glucose deprivation. This methodology not only advances our understanding of subcellular dynamics, but also holds promise for transformative applications in biomedical research and therapeutic development.

Real-Time Monitoring of Tyrosine Hydroxylase Activity with a Ratiometric Fluorescent Probe.

Parkinson's disease (PD) represents the second most widespread neurodegenerative disease, and early monitoring and diagnosis are urgent at present. Tyrosine hydroxylase (TH) is a key enzyme for producing dopamine, the levels of which can serve as an indicator for assessing the severity and progression of PD. This renders the specific detection and visualization of TH a strategically vital way to meet the above demands. However, a fluorescent probe for TH monitoring is still missing. Herein, three rationally designed wash-free ratiometric fluorescent probes were proposed. Among them, TH-1 exhibited ideal photophysical properties and specific dual-channel bioimaging of TH activity in SH-SY5Y nerve cells. Moreover, the probe allowed for in vivo imaging of TH activity in zebrafish brain and living striatal slices of mice. Overall, the ratiometric fluorescent probe TH-1 could serve as a potential tool for real-time monitoring of PD in complex biosystems.

Inflammatory-stimuli-responsive turn-on NIR fluorogenic theranostic prodrug: adjuvant delivery of diclofenac and hydrogen sulfide attenuates acute inflammatory disorders.

Prolonged use of very commonly prescribed non-steroidal anti-inflammatory drugs (NSAIDs) is often associated with undesired side effects, including gastrointestinal ulcers due to the non-selective inhibition of cyclooxygenases. We describe the development of an inflammatory-stimuli-responsive turn-on fluorogenic theranostic prodrug DCF-HS for adjuvant drug delivery. Upon activation by reactive oxygen species (ROS), the prodrug releases diclofenac DCF (active drug) and the NIR fluorophore DCI-NH2 along with carbonyl sulfide (COS). The second activation of COS by the enzyme carbonic anhydrase (CA) generates hydrogen sulfide (H2S). The prodrug was conveniently synthesized using multi-step organic synthesis. The UV-Vis and fluorescence studies revealed the selective reactivity of DCF-HS towards ROS such as H2O2 in the aqueous phase and the desired uncaging of the drug DCF with turn-on NIR fluorescent reporter under physiological conditions. Furthermore, the release of fluorophore DCI-NH2 and drug DCF was confirmed using the reverse phase HPLC method. Compatibility of prodrug activation was studied next in the cellular medium. The prodrug DCF-HS was non-toxic in a representative cancer cell line (HeLa) and a macrophage cell line (RAW 264.7) up to 100 µM concentration, indicating its biocompatibility. The intracellular ROS-mediated activation of the prodrug with the release of NIR dye DCI-NH2 and H2S was investigated in HeLa cells using the H2S-selective probe WSP2. The anti-inflammatory activity of the active drug DCF from the prodrug DCF-HS was studied in the lipopolysaccharide (LPS)-induced macrophage cell line and compared to that of the parent drug DCF using western blot analysis and it was found that the active drug resulted in pronounced inhibition of COX-2 in a dose-dependent manner. Finally, the anti-inflammatory potential of the prodrug and the turn-on fluorescence were validated in the inflammation-induced Wister rat models.

NIR-I emissive cyanine derived molecular probe for selective monitoring of hepatic albumin levels during hyperglycemia.

In this study, we report a small molecule optical marker BI-CyG derived from the structural engineering of a cyanine scaffold. The developed probe offers suitable advantages over existing cyanine-based albumin specific probes in terms of its excitation and emission wavelengths, which are 760 and 830-832 nm, respectively. Structural tuning of the cyanine architecture leading to extended π-conjugation and resulting in a suitable bathochromic shift in the emission wavelength of the probe is represented in this study. The probe besides emitting in the NIR region, also possesses the desirable characteristics of being a potential target selective optical marker, as established from various biophysical studies. Molecular modelling and simulation studies provided critical insights into the binding of the probe in the protein microenvironment, which was further supported by experimental studies. The probe displayed intracellular albumin selectivity and was utilized for demonstrating alteration in albumin levels in pathological states such as hyperglycemia in hepatic cells. The present study also sheds some light on using BI-CyG as an imaging probe and on the role of metformin as a suitable drug for balancing hyperglycemia-induced reduced intra-hepatic albumin levels. The study, thus, attempts to highlight the structural derivatization of cyanine to afford a potential probe for serum albumin and its deployment to image altering albumin levels in an induced pathological condition, hyperglycemia.

A translocation fluorescent probe for analyzing cellular physiological parameters in neurological disease models.

Neurological disorders are closely linked to the alterations in cell membrane permeability (CMP) and mitochondrial membrane potential (MMP). Changes in CMP and MMP may lead to damage and death of nerve cells, thus triggering the onset and progression of neurological diseases. Therefore, monitoring the changes of these two physiological parameters not only benefits the accurate assessment of nerve cell health status, but also enables providing key information for the diagnosis and treatment of neurological diseases. However, the simultaneous monitoring of these two cellular physiological parameters is still challenging. Herein, we design and synthesize two quinolinium-carbazole-derivated fluorescent probes (OQ and PQ). As isomers, the only difference in their chemical structures is the linking position of the carbazole unit in quinoline rings. Strikingly, such a subtle difference endows OQ and PQ with significantly different organelle-staining behaviors. PQ mainly targets at the nucleus, OQ can simultaneously stain cell membranes and mitochondria in normal cells, and performs CMP and MMP-dependent translocation from the cell membrane to mitochondria then to the nucleus, thus holding great promise as an intracellular translocation probe to image the changes of CMP and MMP. After unraveling the intrinsic mechanism of their different translocation abilities by combining experiments with molecular dynamics simulations and density functional theory calculations, we successfully used OQ to monitor the continuous changes of CMP and MMP in three neurological disease-related cell models, including oxidative stress-damaged, Parkinson's disease, and virus-infected ones. Besides providing a validated imaging tool for monitoring cellular physiological parameters, this work paves a promising route for designing intracellular translocation probes to analyze cellular physiological parameters associated with various diseases.

A supramolecular colloidal system based on folate-conjugated ß-cyclodextrin polymer and indocyanine green for enhanced tumor-targeted cell imaging in 2D culture and 3D tumor spheroids.

Indocyanine green (ICG) is an FDA-approved medical diagnostic agent that is widely used as a near-infrared (NIR) fluorescent imaging molecular probe. However, ICG tends to aggregate to form dimers or H-aggregates in water and lacks physical and optical stability, which greatly decreases its absorbance and fluorescence intensity in various applications. Additionally, ICG has no tissue- or tumor-targeting properties, and its structure is not easy to modify, which has further limited its application in cancer diagnosis. In this study, we addressed these challenges by developing a supramolecular colloidal carrier system that targets tumor cells. To this end, we synthesized a water-soluble ß-cyclodextrin (ß-CD) polymer conjugated with folate (FA), denoted PCD-FA, which is capable of forming inclusion complexes with ICG in water through host-guest interactions between the ß-CD moieties and ICG molecules. The inclusion complexes formed by PCD-FA and ICG, called ICG@PCD-FA, dispersed stably in solution as colloidal nanoparticles, greatly improving the physical and optical properties of ICG by preventing ICG dimer formation, where ICG appeared as monomers and even J-aggregates. This resulted in stronger and more stable absorption at a longer wavelength of 900 nm, which may allow for deeper tissue penetration and imaging with reduced interference from biological tissues' autofluorescence. Moreover, ICG@PCD-FA showed a targeting effect on folate receptor-positive (FR+) tumor cells, which specifically highlighted FR+ cells via NIR endoscopic imaging. Notably, ICG@PCD-FA further improved permeation and accumulation in FR+ 3D tumor spheroids. Therefore, this ICG@PCD-FA supramolecular colloidal system may have a great potential for use in tumor NIR imaging and diagnostic applications.

Cascading Detection of Hydrogen Sulfide and N-Acetyltransferase 2 in Hepatocellular Carcinoma Cells Using a Two-Photon Fluorescent Probe.

Hydrogen sulfide (H2S), a critical gas signaling molecule, and N-acetyltransferase 2 (NAT2), a key enzyme in drug metabolism, are both known active biomarkers for liver function. However, the interactions and effects of H2S and NAT2 in living cells or lesion sites remain unknown due to the lack of imaging tools to achieve simultaneous detection of these two substances, making it challenging to implement real-time imaging and precise tracking. Herein, we report an activity-based two-photon fluorescent probe, TPSP-1, for the cascade detection of H2S and NAT2 in living liver cells. Continuous conversion from TPSP-1 to TPSP-3 was achieved in liver cells and tissues. Significantly, leveraging the outstanding optical properties of this two-photon fluorescent probe, TPSP-1, has been effectively used to identify pathological tissue samples directly from clinical liver cancer patients. This work provides us with this novel sensing and two-photon imaging probe, which can be used as a powerful tool to study the physiological functions of H2S and NAT2 and will help facilitate rapid and accurate diagnosis and therapeutic evaluation of hepatocellular carcinoma.

Construction and evaluation of near-infrared fluorescent probes for imaging lipid droplet and lysosomal viscosity.

Microenvironmental viscosity is a crucial parameter for biological systems, and its abnormal fluctuations are closely associated with various functional disorders and diseases. However, it is still important and urgent to develop improved near-infrared fluorescent probes for micro-viscosity with dual-organelle targeting properties, low background noise, and high sensitivity. Herein, two BODIPY-based small-molecule fluorescent probes were designed and synthesized, which were explored for their viscosity- and polarity-responsive properties, and were further applied to imaging sub-cellular viscosity in living cells. Interestingly, BSZ-Ph and BSZ-R displayed near-infrared fluorescence (more than 650 nm) and were sensitive to environmental viscosity and polarity due to the introduction of a benzothiazole at the 2-position and electron-rich aniline groups at the 5-position of the BODIPY core, respectively. The fluorescence intensity increased exponentially with the viscosity changes. Furthermore, the probe BSZ-Ph could successfully target lipid droplets and image cellular viscosity changes by treating lipopolysaccharides (LPS) and nystatin. Comparatively, the probe BSZ-R could successfully target the dual organelles of lipid droplets and lysosomes and image cellular viscosity changes by treating LPS and monensin. Therefore, in this work, we reported two new BODIPY-based near-infrared fluorescent probes, BSZ-Ph and BSZ-R, for cellular viscosity imaging, which could target lipid droplets and the dual organelles of lysosomes and lipid droplets, respectively. The study could provide a reference for the future development of fluorescent probes for viscosity in lipid droplets and lysosomes.

Near-infrared fluorescent indolizine-dicyanomethylene-4H-pyran hybrids for viscosity imaging in living cells.

The development of near-infrared organic fluorescent dyes with tunable emission profiles is highly required in the field of biological sensing and imaging. In this paper, we designed and synthesized two organic fluorescent dyes, DCM-1 and DCM-2, through the hybridization of indolizine and dicyanomethylene-4H-pyran skeleton. These two compounds show near-infrared fluorescence with emission maximum approximately at 640 and 680 nm, respectively. Notably, both DCM-1 and DCM-2 have specific responses to viscosity without being interfered by biological relevant species. Cell experiments demonstrate that DCM-1 and DCM-2 can detect dynamic changes in viscosity within living cells, suggesting their potential applications in chemical biology research.

Design, synthesis and application of dual-channel fluorescent probes for ratiometric detection of HClO and H2S based on phenothiazine coumarins.

The ubiquity of diverse material entities in environmental matrices renders the deployment of unifunctional fluorescent indicators inadequate. Consequently, this study introduces a ratiometric dual-emission fluorescent sensor (Probe CP), synthesized by conjugating phenothiazine coumarin to hydroxycoumarin through a piperazine linker for concurrent detection of HClO and H2S. Upon interaction with HClO, the phenothiazine unit's sulfur atom undergoes oxidation to sulfoxide, facilitating a shift from red to green fluorescence in a ratiometric manner. Concurrently, at the opposite terminus of Probe CP, 2,4-dinitroanisole serves as the reactive moiety for H2S recognition; it restores the blue emission characteristic of 7-hydroxycoumarin while maintaining the red fluorescence emanating from phenothiazine coumarin as an internal standard for ratio-based assessment. Exhibiting elevated specificity and sensitivity coupled with minimal detection thresholds (0.0506 µM for HClO and 1.7292 µM for H2S) alongside rapid equilibration periods (3 min for HClO and half an hour for H2S), this sensor was efficaciously employed in cellular environments and within zebrafish models as well as imaging applications pertaining to alcohol-induced hepatic injury in murine subjects.

Preclinical evaluation of AGTR1-Targeting molecular probe for colorectal cancer imaging in orthotopic and liver metastasis mouse models.

Despite advancements in colorectal cancer (CRC) treatment, the prognosis remains unfavorable for patients with distant liver metastasis. Fluorescence molecular imaging with specific probes is increasingly used to guide CRC surgical resection in real-time and treatment planning. Here, we demonstrate the targeted imaging capacity of an MPA-PEG4-N3-Ang II probe labeled with near-infrared (NIR) fluorescent dye targeting the angiotensin II (Ang II) type 1 receptor (AGTR1) that is significantly upregulated in CRC. MPA-PEG4-N3-Ang II was highly selective and specific to in vitro tumor cells and in vivo tumors in a mouse CRC xenograft model. The favorable ex vivo imaging and in vivo biodistribution of MPA-PEG4-N3-Ang II afforded tumor-specific accumulation with low background and >10 contrast tumor-to-colorectal values in multiple subcutaneous CRC models at 8 h following injection. Biodistribution analysis confirmed the probe's high uptake in HT29 and HCT116 orthotopic and liver metastatic models of CRC with signal-to-noise ratio (SNR) values of tumor-to-colorectal and -liver fluorescence of 5.8 ± 0.6, 5.3 ± 0.7, and 2.7 ± 0.5, 2.6 ± 0.5, respectively, enabling high-contrast intraoperative tumor visualization for surgical navigation. Given its rapid tumor targeting, precise tumor boundary delineation, durable tumor retention and docking study, MPA-PEG4-N3-Ang II is a promising high-contrast imaging agent for the clinical detection of CRC.