Biomimetic NIR-II fluorescent proteins created from chemogenic protein-seeking dyes for multicolor deep-tissue bioimaging.

Near-infrared-I/II fluorescent proteins (NIR-I/II FPs) are crucial for in vivo imaging, yet the current NIR-I/II FPs face challenges including scarcity, the requirement for chromophore maturation, and limited emission wavelengths (typically < 800 nm). Here, we utilize synthetic protein-seeking NIR-II dyes as chromophores, which covalently bind to tag proteins (e.g., human serum albumin, HSA) through a site-specific nucleophilic substitution reaction, thereby creating proof-of-concept biomimetic NIR-II FPs. This chemogenic protein-seeking strategy can be accomplished under gentle physiological conditions without catalysis. Proteomics analysis identifies specific binding site (Cys 477 on DIII). NIR-II FPs significantly enhance chromophore brightness and photostability, while improving biocompatibility, allowing for high-performance NIR-II lymphography and angiography. This strategy is universal and applicable in creating a wide range of spectrally separated NIR-I/II FPs for real-time visualization of multiple biological events. Overall, this straightforward biomimetic approach holds the potential to transform fluorescent protein-based bioimaging and enables in-situ albumin targeting to create NIR-I/II FPs for deep-tissue imaging in live organisms.

Widefield functional speckle-correlation optical scattering mesoscopy toward hemodynamic imaging.

Speckle-correlation optical scattering imaging (SCOSI) has shown the potential for non-invasive biomedical diagnostic applications, which directly utilizes the scattering patterns to reconstruct the deep and non-line-of-sight objects. However, the course of the translation of this technique to preclinical biomedical imaging applications has been postponed by the following two facts: 1) the field of view of SCOSI was significantly limited by the optical memory effect, and 2) the molecular-tagged functional imaging of the biological tissues remains largely unexplored. In this work, a proof-of-concept design of the first-generation widefield functional SCOSI (WF-SCOSI) system was presented for simultaneously achieving mesoscopic mapping of fluid morphology and flow rate, which was realized by implementing the concepts of scanning synthesis and fluorescence scattering flowmetry. The ex vivo imaging results of the fluorescence-labeled large-scale blood vessel network phantom underneath the strong scatters demonstrated the effectiveness of WF-SCOSI toward non-invasive hemodynamic imaging applications.

Biocompatible and Water-Soluble Shortwave-Infrared (SWIR)-Emitting Cyanine-Based Fluorescent Probes for In Vivo Multiplexed Molecular Imaging.

Extending molecular imaging into the shortwave-infrared (SWIR, 900-1400 nm) region provides deep tissue visualization of biomolecules in the living system resulting from the low tissue autofluorescence and scattering. Looking at the Food and Drug Administration-approved and clinical trial near-infrared (NIR) probes, only indocyanine green (ICG) and its analogues have been approved for biomedical applications. Excitation wavelength less than 800 nm limits these probes from deep tissue penetration and noninvasive fluorescence imaging. Herein, we present the synthesis of ICG-based π-conjugation-extended cyanine dyes, ICG-C9 and ICG-C11 as biocompatible, and water-soluble SWIR-emitting probes with emission wavelengths of 922 and 1010 nm in water, respectively. Also, ICG-, ICG-C9-, and ICG-C11-based fluorescent labeling agents have been synthesized for the development of SWIR molecular imaging probes. Using the fluorescence of ICG, ICG-C9, and ICG-C11, we demonstrate three-color SWIR fluorescence imaging of breast tumors by visualizing surface receptors (EGFR and HER2) and tumor vasculature in living mice. Furthermore, we demonstrate two-color SWIR fluorescence imaging of breast tumor apoptosis using an ICG-conjugated anticancer drug, Kadcyla and ICG-C9 or ICG-C11-conjugated annexin V. Finally, we show long-term (38 days) SWIR fluorescence imaging of breast tumor shrinkage induced by Kadcyla. This study provides a general strategy for multiplexed fluorescence molecular imaging with biocompatible and water-soluble SWIR-emitting cyanine probes.

Discovery of Near-Infrared Heptamethine Cyanine Probes for Imaging-Guided Surgery in Solid Tumors.

Near-infrared (NIR) fluorescence imaging has attracted much attention in image-guided interventions with unique advantages. However, the clinical translation rate of fluorescence probes is extremely low, primarily due to weak lesion signal contrast and poor specificity. To address this dilemma, a series of small-molecule near-infrared fluorescence probes have been designed for tumor imaging. Among them, YQ-04-03 showed notable optical stability and remarkable sensitivity toward tumor targeting. Moreover, within a specific concentration and time range against oxidizing reducing agents and laser, it demonstrated better stability than ICG. The retention time of YQ-04-03 in tumors was significantly longer compared to other nonspecific uptake sites in the subjects, and its tumor-to-normal tissue ratio (TNR) outperformed ICG. Successful resection of in situ hepatocarcinoma and peritoneal carcinoma was achieved using probe imaging guidance, with the smallest visual lesion resected measuring approximately 1 mm3. Ultimately, this probe holds great potential for advancing tumor tracer.

ICG-Fluorescence Imaging for Margin Assessment During Minimally Invasive Colorectal Liver Metastasis Resection.

Importance: Unintended tumor-positive resection margins occur frequently during minimally invasive surgery for colorectal liver metastases and potentially negatively influence oncologic outcomes. Objective: To assess whether indocyanine green (ICG)-fluorescence-guided surgery is associated with achieving a higher radical resection rate in minimally invasive colorectal liver metastasis surgery and to assess the accuracy of ICG fluorescence for predicting the resection margin status. Design, Setting, and Participants: The MIMIC (Minimally Invasive, Indocyanine-Guided Metastasectomy in Patients With Colorectal Liver Metastases) trial was designed as a prospective single-arm multicenter cohort study in 8 Dutch liver surgery centers. Patients were scheduled to undergo minimally invasive (laparoscopic or robot-assisted) resections of colorectal liver metastases between September 1, 2018, and June 30, 2021. Exposures: All patients received a single intravenous bolus of 10 mg of ICG 24 hours prior to surgery. During surgery, ICG-fluorescence imaging was used as an adjunct to ultrasonography and regular laparoscopy to guide and assess the resection margin in real time. The ICG-fluorescence imaging was performed during and after liver parenchymal transection to enable real-time assessment of the tumor margin. Absence of ICG fluorescence was favorable both during transection and in the tumor bed directly after resection. Main Outcomes and Measures: The primary outcome measure was the radical (R0) resection rate, defined by the percentage of colorectal liver metastases resected with at least a 1 mm distance between the tumor and resection plane. Secondary outcomes were the accuracy of ICG fluorescence in detecting margin-positive (R1; <1 mm margin) resections and the change in surgical management. Results: In total, 225 patients were enrolled, of whom 201 (116 [57.7%] male; median age, 65 [IQR, 57-72] years) with 316 histologically proven colorectal liver metastases were included in the final analysis. The overall R0 resection rate was 92.4%. Re-resection of ICG-fluorescent tissue in the resection cavity was associated with a 5.0% increase in the R0 percentage (from 87.4% to 92.4%; P < .001). The sensitivity and specificity for real-time resection margin assessment were 60% and 90%, respectively (area under the receiver operating characteristic curve, 0.751; 95% CI, 0.668-0.833), with a positive predictive value of 54% and a negative predictive value of 92%. After training and proctoring of the first procedures, participating centers that were new to the technique had a comparable false-positive rate for predicting R1 resections during the first 10 procedures (odds ratio, 1.36; 95% CI, 0.44-4.24). The ICG-fluorescence imaging was associated with changes in intraoperative surgical management in 56 (27.9%) of the patients. Conclusions and Relevance: In this multicenter prospective cohort study, ICG-fluorescence imaging was associated with an increased rate of tumor margin-negative resection and changes in surgical management in more than one-quarter of the patients. The absence of ICG fluorescence during liver parenchymal transection predicted an R0 resection with 92% accuracy. These results suggest that use of ICG fluorescence may provide real-time feedback of the tumor margin and a higher rate of complete oncologic resection.

[Preliminary application of indocyanine green fluorescence imaging technology in nasal endoscopic tumor surgery].

Objective:To preliminarily study the practical value of Indocyanine green（ICG） molecular fluorescence imaging technology in nasal endoscopic tumor surgery. Methods:Five patients with tumors related to nasal sinuses, orbital wall and skull base in the Department of Otolaryngology head and Neck Surgery, General Hospital of Xinjiang Military Command from December 2022 to April 2023 were enrolled. Among them, 3 were benign tumors and 2 were malignant tumors. All patients underwent surgery under the guidance of ICG molecular fluorescence imaging. ICG was administered intravenously through cubital vein at a dose of 0.5 mg/kg 12 to 24 h before surgery. Tumors were labeled by fluorescence imaging during the operation. surgeons cleared the tumor tissue strictly according to the labeled range and depth, malignant tumors were further expanded and cleaned according to pathology results. Results:All 5 patients achieved accurate tumor localization with the aid of fluorescence imaging technology. Resections were performed with reference to fluorescent labeling boundaries, all patients achieved complete tumor cleanup or negative margins. Conclusion:For tumor-related surgery under nasal endoscopy, ICG molecular fluorescence imaging technology can not only achieve accurate real-time positioning, but also provide evidence for surgeons to judge tumor boundaries. Therefore, we believe that the technology should have certain practical value in nasal endoscopic tumor surgery.

Role of indocyanine green fluorescence imaging for evaluating blood supply in the gastric conduit via the substernal route after McKeown minimally invasive esophagectomy.

BACKGROUND: Anastomotic leakage (AL) is a determining factor of morbidity and mortality after esophagectomy. Adequate perfusion of the gastric conduit is crucial for AL prevention. This study aimed to determine whether intraoperative angiography using indocyanine green (ICG) fluorescence improves the incidence of AL after McKeown minimally invasive esophagectomy (MIE) with gastric conduit via the substernal route (SR). METHODS: This retrospective cohort study included 120 patients who underwent MIE with gastric conduit via SR for esophageal cancer between February 2019 and April 2023. Of 120 patients, 88 experienced intraoperative angiography using ICG (ICG group), and 32 patients experienced intraoperative angiography without ICG (no-ICG group). Baseline characteristics and operative outcomes, including AL as the main concern, were compared between the 2 groups. In addition, the outcomes among patients in the ICG group with different levels of fluorescence intensity were compared. RESULTS: The ICG and no-ICG groups were comparable in baseline characteristics and operative outcomes. There was no significant difference between the 2 groups regarding the rate of AL (31.0% vs 37.5%; P = .505), median dates of AL (9 vs 9 days; P = .810), and severity of AL (88.9%, 11.11%, and 0.0% vs 66.7%, 16.7%, and 16.7% for grades I, II, and III, respectively; P = .074). Patients in the ICG group with lower intensity of ICG had higher rates of leakage (24.6%, 39.3%, and 100% in levels I, II, and III of ICG intensity, respectively; P = .04). CONCLUSION: The use of ICG did not seem to reduce the rate of AL. However, abnormal intensity of ICG fluorescence was associated with a higher rate of AL, which implies a predictive potential.

De Novo Design of a Highly Stable Ratiometric Probe for Long-Term Continuous Imaging of Endogenous HClO Burst.

Long-term continuous imaging of endogenous HClO burst is of great importance for the elucidation of various physiological or pathological processes. However, most of the currently reported HClO probes have failed to achieve this goal due to their insufficient photobleaching resistance under a laser source. Herein, a highly stable ratiometric probe, HFTC-HClO 1, which is capable of continuously monitoring endogenous HClO burst over a long period of time, has been judiciously developed. Briefly, the de novo development of HFTC-HClO 1 mainly involved three main steps: (1) novel coumarins (HFTC 1-5) were designed and synthesized; (2) the most stable scaffold, HFTC 3, was selected through dye screening and cell imaging validation; and (3) based on HFTC 3, three candidate HClO probes were constructed, and HFTC-HClO 1 was finally selected due to its superior sensing properties toward HClO. Furthermore, HFTC-HClO 1 can quantitatively measure HClO levels in various real samples with excellent recovery (>90.4%), and the use of HFTC-HClO 1-coated test strips for qualitative analysis of HClO in real samples was also achieved. In addition, the application of HFTC-HClO 1 for long-term continuous monitoring of intracellular HClO burst was successfully demonstrated. Significantly, HFTC-HClO 1 was able to visualize HClO generated in the rheumatoid arthritis mouse model.

Molecular Engineering of Activatable NIR-II Hemicyanine Reporters for Early Diagnosis and Prognostic Assessment of Inflammatory Bowel Disease.

Molecular imaging in the second near-infrared window (NIR-II) provides high-fidelity visualization of biopathological events in deep tissue. However, most NIR-II probes produce "always-on" output and demonstrate poor signal specificity toward biomarkers. Herein, we report a series of hemicyanine reporters (HBCs) with tunable emission to NIR-II window (715-1188 nm) and structurally amenable to constructing activatable probes. Such manipulation of emission wavelengths relies on rational molecular engineering by integrating benz[c,d]indolium, benzo[b]xanthonium, and thiophene moieties to a conventional hemicyanine skeleton. In particular, HBC4 and HBC5 possess bright and record long emission over 1050 nm, enabling improved tissue penetration depth and superior signal to background ratio for intestinal tract mapping than NIR-I fluorophore HC1. An activatable inflammatory reporter (AIR-PE) is further constructed for pH-triggered site-specific release in colon. Due to minimized background interference, oral gavage of AIR-PE allows clear delineation of irritated intestines and assessment of therapeutic responses in a mouse model of inflammatory bowel disease (IBD) through real-time NIRF-II imaging. Benefiting from its high fecal clearance efficiency (>90%), AIR-PE can also detect IBD and evaluate the effectiveness of colitis treatments via in vitro optical fecalysis, which outperforms typical clinical assays including fecal occult blood testing and histological examination. This study thus presents NIR-II molecular scaffolds that are not only applicable to developing versatile activatable probes for early diagnosis and prognostic monitoring of deeply seated diseases but also hold promise for future clinical translations.

Application of Laennec extrathecal blockade combined with indocyanine green fluorescence imaging in laparoscopic anatomic hepatectomy.

OBJECTIVE: To investigate the safety and application value of combining Laennec extracapsular occlusion with ICG fluorescence imaging in laparoscopic anatomic hepatectomy. METHODS: Complete laparoscopic dissection was performed outside the Laennec sheath, blocking Glisson's pedicle of the corresponding liver segment or lobe. An appropriate amount of indocyanine green (ICG) dye was intravenously injected, and the boundary line between the pre-cut liver segment and liver lobe was identified using fluorescence laparoscopy. Complete resection of the liver segment or lobe was performed based on anatomical markers. Clinical data, including operation time, intraoperative blood loss, postoperative hospital stay, and postoperative complications, were collected. RESULTS: A total of 14 cases were included in the study, including seven cases of primary liver cancer, three cases of metastatic liver cancer, three cases of intrahepatic bile duct calculi, and one case of hepatic hemangioma. All 14 patients underwent anatomic hepatectomy under fluorescent laparoscopy, with four cases involving the right liver, seven cases involving the left liver, two cases involving the right anterior lobe, and one case involving the right posterior lobe. CONCLUSION: Combining laparoscopic follow-up of the Laennec membrane with Glisson outer sheath block and intraoperative ICG fluorescence imaging provides real-time guidance for locating the resection boundaries during anatomic hepatectomy. This approach helps in controlling intraoperative bleeding, reducing operation time, and ensuring high safety. It holds significant value in clinical application.

Natural Fat Nanoemulsions for Enhanced Optical Coherence Tomography Neuroimaging and Tumor Imaging in the Second Near-Infrared Window.

Optical coherence tomography (OCT) imaging mainly uses backscattered light to visualize the structural and functional information on biological tissues. In particular, OCT angiography can not only map the capillary networks but also capture the blood flow in the tissue microenvironment, making it a good candidate for neuroimaging and tumor imaging in vivo and in real time. To further improve the detection accuracy of cancer or brain disorders, it is essential to develop a natural and nontoxic contrast agent for enhanced OCT imaging in the second near-infrared (NIR-II) window. In this study, a superior biocompatible and highly scattering NIR-II fat nanoemulsion was constructed to improve OCT imaging contrast and depth for monitoring the vascular network changes of the cerebral cortex or tumor. In vivo experimental results demonstrated that a natural fat nanoemulsion can serve as an excellent probe for enhanced OCT neuroimaging and tumor imaging.

Development of a turn-on fluorescent probe for the imaging of intracellular hypochlorous acid (HClO) during ferroptosis.

Ferroptosis is a burgeoning iron-dependent cell death form, and has close relation with hypochlorous acid (HClO). Exploring the fluctuation of the HClO level in living cells during ferroptosis could contribute to the profound study of the biological functions of HClO during ferroptosis. Here, we present a turn-on probe (RH-C) for the imaging of intracellular HClO during ferroptosis. The probe RH-C utilized the N,N-dimethylthiocarbamate group as a selective recognition site for HClO, and displayed desirable sensitivity and selectivity to HClO. The probe RH-C could detect the exogenous and endogenous HClO in living cells. Furthermore, RH-C was competent in monitoring the changes of endogenous HClO level during the process of ferroptosis. Biological imaging results suggested that erastin-induced ferroptosis can result in the excessive production of the endogenous HClO, and ferrostatin-1 (Fer-1) and vitamin E (VE) could block the massive accumulation of HClO in living cells.

A pyrene-induced PET-based chemosensor for biologically important Zn(II) ions: application in test strips and live cell imaging studies.

Cation and anion sensing is vital owing to their universal dispersion in ecosystems and biological functions. It has been shown that fluorescent receptors based on organic platforms are efficient for detecting a number of ions and have many advantages such as low cost, superior sensitivity and simplicity in installation. This study demonstrates the design and synthesis of a novel receptor (E)-3-[(3,5-di-tert-butyl-2-hydroxybenzylidene)amino]-2-(pyren-1-yl)-2,3-dihydroquinazolin-4(1H)-one (DTQ) for the rapid recognition of Zn(II) ions. DTQ exhibited a significant fluorometric "turn-on" characteristic towards Zn(II) at λmax 444 nm in aqueous acetonitrile by inhibiting the photo-induced electron transfer (PET) and -CN- process. The ESI-MS analysis and Job's plot experimental results confirmed stoichiometric 1 : 1 complex formation between DTQ and Zn(II). Fluorometric investigations revealed the detection limit and association constant of DTQ towards Zn(II), which were found to be 13.4 nM and 1.47 × 105 M-1, respectively. DTQ was employed to sense Zn(II) on low-cost test strips. The present research findings imply that DTQ can function as an effective sensor for Zn(II).

Near-Infrared Fluorescence Imaging Sensor with Laser Diffuser for Visualizing Photoimmunotherapy Effects under Endoscopy.

The drug efficacy evaluation of tumor-selective photosensitive substances was expected to be enabled by imaging the fluorescence intensity in the tumor area. However, fluorescence observation is difficult during treatments that are performed during gastrointestinal endoscopy because of the challenges associated with including the fluorescence filter in the camera part. To address this issue, this study developed a device that integrates a narrow camera and a laser diffuser to enable fluorescence imaging through a forceps port. This device was employed to demonstrate that a laser diffuser with an NIR fluorescence imaging sensor could be delivered through a 3.2 mm diameter port. In addition, fluorescence images of Cetuximab-IR700 were successfully observed in two mice, and the fluorescence intensity confirmed that the fluorescence decayed within 330 s. This device is expected to have practical application as a tool to identify the optimal irradiation dose for tumor-selective photosensitive substances under endoscopy.

JS-K Combined with a Melanin-Based Theranostic Agent: A Novel Sequential Delivery Strategy to Enhance the Near-Infrared Fluorescence Imaging of Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a 5 year survival rate less than 12%. This malignancy is closely related to the unique tumor microenvironment (TME), which is characterized by a hypovascular and hyperdense extracellular matrix, making it difficult for drugs to permeate the tumor center. Near-infrared fluorescence (NIRF) imaging, which has high sensitivity and resolution, may improve the survival rate of PDAC patients. In this study, we first used JS-K (O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl) piperazine-1-yl] diazene-1-ium-1,2-diolate) to specifically dilate blood vessels within the TME of PDAC patients and subsequently injected IR820-PEG-MNPs (IPM NPs) to diagnose and treat orthotopic PDAC. We found that JS-K promoted the accumulation of IPM NPs in orthotopic Pan02 tumor-bearing mice and was able to increase the tumor signal-to-background ratio (SBR) in the orthotopic PDAC area by 41.5%. In addition, surgical navigation in orthotopic Pan02 tumor-bearing mice and complete tumor resection based on fluorescence imaging were achieved with a detection sensitivity of 81.0%. Moreover, we verified the feasibility of the combination of laparoscopy and photothermal ablation (PTA) for the treatment of PDAC. Finally, we demonstrated that IPM NPs had greater affinity for human PDAC tissues than for normal pancreatic tissues ex vivo, preliminarily highlighting the potential for clinical translation of these NPs. In conclusion, we developed and validated a novel sequential delivery strategy that promotes the accumulation of nanoagents in the tumor area and can be used for the diagnosis and treatment of PDAC.

Site-specific albumin tagging with NIR-II fluorogenic dye for high-performance and super-stable bioimaging.

Synthetic near-infrared-II (NIR-II) dyes are promising for deep tissue imaging, yet they are generally difficult to target a given biomolecule with high specificity. Furthermore, the interaction mechanism between albumin and cyanine molecules, which is usually regarded as uncertain "complexes" such as crosslinked nanoparticles, remains poorly understood. Methods: Here, we propose a new class of NIR-II fluorogenic dyes capable of site-specific albumin tagging for in situ albumin seeking/targeting or constructing high-performance cyanine@albumin probes. We further investigate the interaction mechanism between NIR-II fluorogenic dyes and albumin. Results: We identify CO-1080 as an optimal dye structure that produces a stable/bright NIR-II cyanine@albumin probe. CO-1080 exhibits maximum supramolecular binding affinity to albumin while catalyzing their covalent attachment. The probe shows exact binding sites located on Cys476 and Cys101, as identified by proteomic analysis and docking modeling. Conclusion: Our cyanine@albumin probe substantially improves the pharmacokinetics of its free dye counterpart, enabling high-performance NIR-II angiography and lymphography. Importantly, the site-specific labeling tags between NIR-II fluorogenic dyes and albumin occur under mild conditions, offering a specific and straightforward synthesis strategy for NIR-II fluorophores in the fields of targeting bioimaging and imaging-guided surgery.

Activatable Dual-Optical Molecular Probe for Bioimaging Superoxide Anion in Epilepsy.

Superoxide anion (O2•-) plays a pivotal role in the generation of other reactive oxygen species within the body and is closely linked to epilepsy. Despite this connection, achieving precise imaging of O2•- during epilepsy pathology remains a formidable challenge. Herein, we develop an activatable molecular probe, CL-SA, to track the fluctuation of the level of O2•- in epilepsy through simultaneous fluorescence imaging and chemiluminescence sensing. The developed probe CL-SA demonstrated its efficacy in imaging of O2•- in neuronal cells, showcasing its dual optical imaging capability for O2•- in vitro. Furthermore, CL-SA was successfully used to observe aberrantly expressed O2•- in a mouse model of epilepsy. Overall, CL-SA provides us with a valuable tool for chemical and biomedical studies of O2•-, promoting the investigation of O2•- fluctuations in epilepsy, as well as providing a reliable means to explore the diagnosis and therapy of epilepsy.

Live Cell Imaging and Real-Time Monitoring of Nucleolus Morphology and Mitophagy with a Red Fluorescent and Photostable rRNA-Specific Probe in Human Cancer Cells.

rRNAs are prevalent in living organisms. They are produced in nucleolus and mitochondria and play essential cellular functions. In addition to the primary biofunction in protein synthesis, rRNAs have been recognized as the emerging signaling molecule and drug target for studies on nucleolus morphology, mitochondrial autophagy, and tumor cell malignancy. Currently, only a few rRNA-selective probes have been developed, and most of them encounter the drawbacks of low water solubility, poor nuclear membrane permeability, short emission wavelength, low stability against photobleaching, and high cytotoxicity. These unfavorable properties of rRNA probes limit their potential applications. In the present study, we reported a new rRNA-selective and near-infrared fluorescent turn-on probe, 4MPS-TO, capable of tracking rRNA in live human cancer cells. The real-time monitoring performance in nucleolus morphology and mitochondrial autophagy is demonstrated in HeLa cells. The probe shows great application potential for being used as a rRNA-selective, sensitive, and photostable imaging tool in chemical biology study and drug screening.

An automatic parathyroid recognition and segmentation model based on deep learning of near-infrared autofluorescence imaging.

INTRODUCTION: Near-infrared autofluorescence imaging (NIFI) can be used to identify parathyroid gland (PG) during surgery. The purpose of the study is to establish a new model, help surgeons better identify, and protect PGs. METHODS: Five hundred and twenty three NIFI images were selected. The PGs were recorded by NIFI and marked with artificial intelligence (AI) model. The recognition rate for PGs was calculated. Analyze the differences between surgeons of different years of experience and AI recognition, and evaluate the diagnostic and therapeutic efficacy of AI model. RESULTS: Our model achieved 83.5% precision and 57.8% recall in the internal validation set. The visual recognition rate of AI model was 85.2% and 82.4% on internal and external sets. The PG recognition rate of AI model is higher than that of junior surgeons (p < 0.05). CONCLUSIONS: This AI model will help surgeons identify PGs, and develop their learning ability and self-confidence.