Albumin-seeking dyes with adjustable assemblies in situ enable programmable imaging windows and targeting tumor imaging.

Cyanine dyes are widely used organic probes for in vivo imaging due to their tunable fluorescence. They can form complexes with endogenous albumin, resulting in enhanced brightness and photostability. However, this binding is uncontrollable and irreversible, leading to considerable nonspecific background signals and unregulated circulation time. Methods: Here, we connect varying numbers of 4-(4-iodophenyl) butanoic acid (IP) as albumin-binding moieties (ABM) to the cyanine dye, enabling dynamic and controllable binding with albumin. Meanwhile, we provide a blocking method to completely release the dye from covalent capture with albumin, resulting in specific targeting fluorescence. Furthermore, we evaluate the pharmacokinetics and tumor targeting of the developed dyes. Results: The engineered dyes can dynamically and selectively bind with multiple albumins to change the in situ size of assemblies and circulation time, providing programmable regulation over the imaging time window. The nucleophilic substitution of meso-Cl with water-soluble amino acids or targeting peptides for IP-engineered dye further addresses the nonspecific signals caused by albumin, allowing for adjustable angiography time and efficient tumor targeting. Conclusion: This study rationalizes the binding modes of dyes and proteins, applicable to a wide range of near-infrared (NIR) dyes for improving their in vivo molecular imaging.

Near-Infrared-II Imaging Revealed Hypothermia Regulates Neuroinflammation Following Brain Injury by Increasing the Glymphatic Influx.

Advanced in vivo imaging techniques have facilitated the comprehensive visual exploration of animal biological processes, leading to groundbreaking discoveries such as the glymphatic system. However, current limitations of macroscopic imaging techniques impede the precise investigation of physiological parameters regulating this specialized lymphatic transport system. While NIR-II fluorescence imaging has demonstrated advantages in peripheral lymphatic imaging, there are few reports regarding its utilization in the glymphatic system. To address this, a noninvasive transcranial macroscopic NIR-II fluorescence imaging model is developed using a cyanine dye-protein coupled nanoprobe. NIR-II imaging with high temporal and spatial resolution reveals that hypothermia can increase the glymphatic influx by reducing the flow rate of cerebrospinal fluid. In addition, respiratory rate, respiratory amplitude, and heart rate all play a role in regulating the glymphatic influx. Thus, targeting the glymphatic influx may alter the trajectory of immune inflammation following brain injury, providing therapeutic prospects for treating brain injury with mild hypothermia.

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.

An ultra-small organic dye nanocluster for enhancing NIR-II imaging-guided surgery outcomes.

PURPOSE: The accuracy of surgery for patients with solid tumors can be greatly improved through fluorescence-guided surgery (FGS). However, existing FGS technologies have limitations due to their low penetration depth and sensitivity/selectivity, which are particularly prevalent in the relatively short imaging window (< 900 nm). A solution to these issues is near-infrared-II (NIR-II) FGS, which benefits from low autofluorescence and scattering under the long imaging window (> 900 nm). However, the inherent self-assembly of organic dyes has led to high accumulation in main organs, resulting in significant background signals and potential long-term toxicity. METHODS: We rationalize the donor structure of donor-acceptor-donor-based dyes to control the self-assembly process to form an ultra-small dye nanocluster, thus facilitating renal excretion and minimizing background signals. RESULTS: Our dye nanocluster can not only show clear vessel imaging, tumor and tumor sentinel lymph nodes definition, but also achieve high-performance NIR-II imaging-guided surgery of tumor-positive sentinel lymph nodes. CONCLUSION: In summary, our study demonstrates that the dye nanocluster-based NIR-II FGS has substantially improved outcomes for radical lymphadenectomy.

NIR-II nanoprobes for investigating the glymphatic system function under anesthesia and stroke injury.

The glymphatic system plays an important role in the transportation of cerebrospinal fluid (CSF) and the clearance of metabolite waste in brain. However, current imaging modalities for studying the glymphatic system are limited. Herein, we apply NIR-II nanoprobes with non-invasive and high-contrast advantages to comprehensively explore the function of glymphatic system in mice under anesthesia and cerebral ischemia-reperfusion injury conditions. Our results show that the supplement drug dexmedetomidine (Dex) enhances CSF influx in the brain, decreases its outflow to mandibular lymph nodes, and leads to significant differences in CSF accumulation pattern in the spine compared to isoflurane (ISO) alone, while both ISO and Dex do not affect the clearance of tracer-filled CSF into blood circulation. Notably, we confirm the compromised glymphatic function after cerebral ischemia-reperfusion injury, leading to impaired glymphatic influx and reduced glymphatic efflux. This technique has great potential to elucidate the underlying mechanisms between the glymphatic system and central nervous system diseases.

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.

NIR-II Ratiometric Fluorescence Probes Enable Precise Determination of the Metastatic Status of Sentinel Lymph Nodes.

Accurately determining the metastatic status of sentinel lymph nodes (SLNs) through noninvasive imaging with high imaging resolution and sensitivity is crucial for cancer therapy. Herein, we report a dual-tracer-based NIR-II ratiometric fluorescence nanoplatform combining targeted and nontargeted moieties to determine the metastatic status of SLNs through the recording of ratio signals. Ratiometric fluorescence imaging revealed approximately 2-fold increases in signals in tumor-draining SLNs compared to inflamed and normal SLNs. Additionally, inflamed SLNs were diagnosed by combining the ratio value with the enlarged size outputted by NIR-II fluorescence imaging. The metastatic status diagnostic results obtained through NIR-II ratiometric fluorescence signals were further confirmed by standard H&E staining, indicating that the ratiometric fluorescence strategy could achieve distant metastases detection. Furthermore, the superior imaging quality of ratiometric probes enables visualization of the detailed change in the lymphatic network accompanying tumor growth. Compared to clinically available and state-of-the-art NIR contrast agents, our dual-tracer-based NIR-II ratiometric fluorescence probes provide significantly improved performance, allowing for the quick assessment of lymphatic function and guiding the removal of tumor-infiltrating SLNs during cancer surgery.

NIR-II Protein-Escaping Dyes Enable High-Contrast and Long-Term Prognosis Evaluation of Flap Transplantation.

Real-time vascular positioning, postoperative flap monitoring, and vascular reconstruction assessment are of great importance in flap transplantation. Cyanine dyes offer the advantage of high resolution in the Near-infrared-II (NIR-II) imaging window. However, the nonspecific binding of many cyanine dyes to endogenous albumin leads to high organ accumulation and skin absorption, resulting in low-quality imaging and poor reproducibility of contrast during long-term (e.g., 7 days) postoperative monitoring. Here, a novel strategy is proposed that can be widely applied to prevent protein binding for NIR-I/II Cl-containing cyanine dyes. This strategy produces protein-escaping dyes, ensuring high fluorescence enhancement in the blood with rapid clearance and no residual fluorescence, allowing for short-term repeatable injections for vascular imaging. This strategy in the perioperative monitoring of pedicle perforator flap models in mice and rats is successfully applied. Furthermore, leveraging the universality of this strategy, multiple nonoverlapping protein-escaping probes that achieve dual-excitation (808 and 1064 nm) interference-free imaging of nerve-vessel and tumor-vessel simultaneously are designed and synthesized. These protein-escaping dyes enable long-term repeatable dual-color imaging of tumor localization, resection, and tumor-vessel reconstruction at the wound site.

Naked-Eye Readable Microarray for Rapid Profiling of Antibodies against Multiple SARS-CoV-2 Variants.

Novel high-throughput protein detection technologies are critically needed for population-based large-scale SARS-CoV-2 antibody detection as well as for monitoring quality and duration of immunity against virus variants. Current protein microarray techniques rely heavily on labeled transduction methods that require sophisticated instruments and complex operations, limiting their clinical potential, particularly for point-of-care (POC) applications. Here, we developed a label-free and naked-eye readable microarray (NRM) based on a thickness-sensing plasmon ruler, enabling antibody profiling within 30 min. The NRM chips provide 100% accuracy for neutralizing antibody detection by efficiently screening antigen types and experimental conditions and allow for the profiling of antibodies against multiple SARS-CoV-2 variants in clinical samples. We further established a flexible "barcode" NRM assay with a simple tape-based operation, enabling an effective smartphone-based readout and analysis. These results demonstrate new strategies for high-throughput protein detection and highlight the potential of novel protein microarray techniques for realistic clinical applications.

High-precision detection and navigation surgery of colorectal cancer micrometastases.

Surgical resection is an effective treatment for colorectal cancer (CRC) patients, whereas occult metastases hinder the curative effect. Currently, there is no effective method to achieve intraoperatively diagnosis of tumor-positive lymph nodes (LNs). Herein, we adopt a near-infrared-II (NIR-II) organic donor-pi-acceptor-pi-donor probe FE-2PEG, which exhibits bright fluorescence over 1100 nm, excellent photostability, blood circulation time, and biocompatibility, to achieve high-performance bioimaging with improved temporal and spatial resolution. Importantly, the FE-2PEG shows efficient passive enrichment in orthotopic CRC, metastatic mesenteric LNs, and peritoneal metastases by enhanced permeability and retention effect. Under NIR-II fluorescence-guided surgery (FGS), the peritoneal micrometastases were resected with a sensitivity of 94.51%, specificity of 86.59%, positive predictive value (PPV) of 96.57%, and negative predictive value of 79.78%. The PPV still achieves 96.07% even for micrometastases less than 3 mm. Pathological staining and NIR-II microscopy imaging proved that FE-2PEG could successfully delineate the boundary between the tumor and normal tissues. Dual-color NIR-II imaging strategy with FE-2PEG (1100 ~ 1300 nm) and PbS@CdS quantum dots (> 1500 nm) successfully protects both blood supply and normal tissues during surgery. The NIR-II-based FGS provides a promising prospect for precise intraoperative diagnosis and minimally invasive surgery of CRC.

Fe(III)-Shikonin supramolecular nanomedicines as immunogenic cell death stimulants and multifunctional immunoadjuvants for tumor vaccination.

Immunoadjuvants, as an indispensable component of tumor vaccines, can observably enhance the magnitude, breadth, and durability of antitumor immunity. However, current immunoadjuvants suffer from different issues such as weak immunogenicity, inadequate cellular internalization, poor circulation time, and mono-functional bioactivity. Methods: Herein, we construct Fe3+-Shikonin metal-phenolic networks (FeShik) nanomedicines as immunogenic cell death (ICD) stimulants and multifunctional immunoadjuvants for tumor vaccination. The multifunctionality of FeShik nanomedicines is investigated by loading ovalbumin (OVA) as the model antigen to construct OVA@FeShik nanovaccines or 4T1 tumor cell fragment (TF) as homologous antigen to construct TF@FeShik nanovaccines. In vitro examinations including GSH responsive, •OH generation, colloid stability, cellular uptake, cytotoxicity mechanism of ferroptosis and necroptosis, ICD effect, the promotion of DC maturation and antigen cross-presentation were studied. In vivo observations including pharmacokinetics and biodistribution, antitumor effect, abscopal effect, immune memory effect, and biosafety were performed. Results: The presence of FeShik nanomedicines can significantly prolong the blood circulation time of antigens, increasing the bioavailability of antigens. Upon phagocytosis by tumor cells, FeShik nanomedicines can disassemble into Fe2+ and Shikonin in response to tumor microenvironments, leading to ICD of tumor cells via ferroptosis and necroptosis. Consequently, ICD-released autologous tumor cell lysates and pro-inflammatory cytokines not only stimulate DC maturation and antigen cross-presentation, but also promote macrophage repolarization and cytotoxic T lymphocyte infiltration, resulting in the activation of adaptive immune responses toward solid tumors. Conclusion: In a word, our FeShik supramolecular nanomedicines integrate bioactivities of ICD stimulants and immunoadjuvants, such as eradicating tumor cells, activating antitumor immune responses, modulating immunosuppressive tumor microenvironments, and biodegradation after immunotherapy. Encouraged by the diversity of polyphenols and metal ions, our research may provide a valuable paradigm to establish a large library for tumor vaccination.

Dye-Triplet-Sensitized Downshifting Nanoprobes with Ratiometric Dual-NIR-IIb Emission for Accurate In Vivo Detection.

Despite the emerging near-infrared-IIb (NIR-IIb, 1500-1700 nm) bioimaging significantly improving the in vivo penetration depth and resolution, quantitative detection with accuracy remains challenging due to its inhomogeneous fluorescence signal attenuation in biological tissue. Here, ratiometric dual-NIR-IIb in vivo detection with excitation wavelengths of 808 and 980 nm is presented using analyte-responsive dye-triplet-sensitized downshifting nanoprobes (DSNPs). NIR cyanine dye IR-808, a recognizer of biomarker hypochlorite (ClO-), is introduced to trigger a triplet energy transfer process from the dye to Er3+ ions of DSNPs under 808 nm excitation, facilitating the formation of an analyte-responsive 1525 nm NIR-IIb assay channel. Meanwhile, DSNPs also enable emitting intrinsic nonanalyte-dependent downshifting fluorescence at the same NIR-IIb window under 980 nm excitation, serving as a self-calibrated signal to alleviate the interference from the probe amount and depth. Due to the two detected emissions sharing identical light propagation and scattering, the ratiometric NIR-IIb signal is demonstrated to ignore the depth of penetration in biotissue. The arthritis lesions are distinguished from normal tissue using ratiometric probes, and the amount of ClO- can be accurately output by the established detection curves.

Site-specific albumin tagging with chloride-containing near-infrared cyanine dyes: molecular engineering, mechanism, and imaging applications.

Near-infrared dyes, particularly cyanine dyes, have shown great potential in biomedical imaging due to their deep tissue penetration, high resolution, and minimal tissue autofluorescence/scattering. These dyes can be adjusted in terms of absorption and emission wavelengths by modifying their chemical structures. The current issues with cyanine dyes include aggregation-induced quenching, poor photostability, and short in vivo circulation time. Encapsulating cyanine dyes with albumin, whether exogenous or endogenous, has been proven to be an effective strategy for improving their brightness and pharmacokinetics. In detail, the chloride-containing (Cl-containing) cyanine dyes have been found to selectively bind to albumin to achieve site-specific albumin tagging, resulting in enhanced optical properties and improved biosafety. This feature article provides an overview of the progress in the covalent binding of Cl-containing cyanine dyes with albumin, including molecular engineering methods, binding sites, and the selective binding mechanism. The improved optical properties of cyanine dyes and albumin complexes have led to cutting-edge applications in biological imaging, such as tumor imaging (diagnostics) and imaging-guided surgery.

Ultra-Bright Heptamethine Dye Clusters Based on a Self-Adaptive Co-Assembly Strategy for NIR-IIb Biomedical Imaging.

Despite the wide range of applications of bright NIR-II polymethine scaffolds in biomedical imaging, their solvatochromism and aggregation-caused quenching (ACQ) effects in aqueous solutions limit their inherent brightness using traditional encapsulation methods, and effective hydrophilization strategies are still scarce. Here, a new set of Flav dyes is synthesized and PEGylated, followed by manufacturing DSPE@FlavP2000 nanoparticles using a self-adaptive co-assembly strategy to overcome these limitations. FlavP2000 can autonomously adjust its conformation when co-assembled with DSPE-PEG2000 , resulting in high-efficiency luminescence (≈44.9% fluorescence of Flav in DMSO). DSPE@FlavP2000 enables NIR-IIb (>1500 nm) angiography with high signal-to-noise ratios. Notably, this co-assembly can occur in situ between FlavP2000 with proteins in the living body based on a novel mechanism of brightness activation induced by disassembly (BAD), achieving consistent brightness as DSPE@FlavP2000 in blood or serum. The self-adaptive co-assembly strategy can be enhanced by incorporating an IPA moiety, which dynamically binds to albumin to prolong the dye's blood circulation time. Thus, the "enhanced" BAD is successfully applied to long-term vascular imaging and sciatic nerve imaging. Both the self-adaptive co-assembly strategy and BAD phenomenon improve the selectivity and availability of the hydrophilization methods, paving the way for efficient biological applications of polymethine dyes.

Three-Dimensional Histological Electrophoresis for High-Throughput Cancer Margin Detection in Multiple Types of Tumor Specimens.

Accurate identification of tumor margins during cancer surgeries relies on a rapid detection technique that can perform high-throughput detection of multiple suspected tumor lesions at the same time. Unfortunately, the conventional histopathological analysis of frozen tissue sections, which is considered the gold standard, often demonstrates considerable variability, especially in many regions without adequate access to trained pathologists. Therefore, there is a clinical need for a multitumor-suitable complementary tool that can accurately and high-throughput assess tumor margins in every direction within the surgically resected tissue. We herein describe a high-throughput three-dimensional (3D) histological electrophoresis device that uses tumor-specific proteins to identify and contour tumor margins intraoperatively. Testing on seven cell-line xenograft models and human cervical cancer models (representing five types of tissues) demonstrated the high-throughput detection utility of this approach. We anticipate that the 3D histological electrophoresis device will improve the accuracy and efficiency of diagnosing a wide range of cancers.

Three-dimensional histological electrophoresis enables fast automatic distinguishment of cancer margins and lymph node metastases.

Tissue diagnosis is important during surgical excision of solid tumors for margin evaluation. Conventional histopathologic methods rely heavily on image-based visual diagnosis by specialized pathologists, which can be time-consuming and subjective. We report a three-dimensional (3D) histological electrophoresis system for rapid labeling and separation of the proteins within tissue sections, providing a more precise assessment of tumor-positive margin in surgically resected tissues. The 3D histological electrophoresis system uses a tumor-seeking dye labeling strategy to visualize the distribution of tumor-specific proteins within sections and a tumor finder that automatically predicts the tumor contour. We successfully demonstrated the system's capability to predict the tumor contours from five murine xenograft models and distinguish the tumor-invaded region of sentinel lymph nodes. Specifically, we used the system to accurately assess tumor-positive margins from 14 patients with cancer. Our 3D histological electrophoresis system serves as an intraoperative tissue assessment technology for more accurate and automatic pathologic diagnosis.

Protein@Cyanine-Based NIR-II Lymphography Enables the Supersensitive Visualization of Lymphedema and Tumor Lymphatic Metastasis.

Visualization of the lymphatic system is clinically indispensable for the diagnosis and/or treatment of lymphatic diseases. Although indocyanine green (ICG) lymphography becomes an alternate imaging modality compared to traditional lymphoscintigraphy, it is still far from ideal due to the insufficient detection depth and low spatiotemporal resolution. Herein, protein@cyanine probes are rationally developed to solve the limitations of the current near-infrared-I (NIR-I) lymphography. The protein@cyanine probes are synthesized following a chlorine-containing dye-labeling strategy based on structure-selectivity (facile covalent binding between the dye and protein with a 1:1 molar ratio). As expected, the probes display exceptional NIR-II imaging ability with much-improved imaging contrast/resolution and controllable pharmacokinetics, superior to the clinical ICG. The protein@cyanine probes locate lymph nodes and delineate lymphatic vessels with super-high sensitivity and signal-to-background ratio, enabling real-time diagnosing lymphatic diseases such as lymphedema and tumor lymphatic metastasis. In particular, the NIR-II lymphography provides an opportunity to discover the disparate morbidity rate of primary lymphedema in different types of mice. Given the fact of lacking clinically transferable NIR-II probes, this work not only provides a promising strategy for enriching of the current library of NIR-II probes, but also promotes the clinical translation of NIR-II lymphography technology.

Furan Donor for NIR-II Molecular Fluorophores with Enhanced Bioimaging Performance.

The second near-infrared (NIR-II, 1,000 to 1,700 nm) molecular fluorophores containing donor-acceptor-donor conjugated backbone have attracted substantial attention due to their outstanding advantages, such as stable emission and facilely tuned photophysical properties. However, it is still challenging for them to simultaneously achieve high brightness and red-shifted absorption and emission. Herein, furan is adopted as the D unit to construct NIR-II fluorophores, demonstrating red shift of absorption, enhanced absorption coefficient, and fluorescent quantum yield when compared with the generally used thiophene counterparts. The high brightness and desirable pharmacokinetics of the optimized fluorophore, IR-FFCHP, endows improved performance for angiography and tumor-targeting imaging. Furthermore, dual-NIR-II imaging of tumor and sentinel lymph nodes (LNs) has been achieved with IR-FFCHP and PbS/CdS quantum dots, enabling the in vivo imaging navigated LN surgery in tumor-bearing mice. This work demonstrates the potential of furan for constructing bright NIR-II fluorophores for biological imaging.

Exploiting synergistic effect of CO/NO gases for soft tissue transplantation using a hydrogel patch.

Autologous skin flap transplantation is a common method for repairing complex soft tissue defects caused by cancer, trauma, and congenital malformations. Limited blood supply range and post-transplantation ischemia-reperfusion injury can lead to distal necrosis of the flap and long-term functional loss, which severely restricts the decision-making regarding the optimal surgical plan. To address this issue, we develop a hydrogel patch that releases carbon monoxide and nitric oxide gases on demand, to afford a timely blood supply for skin flap transplantation during surgery. Using an ischemia-reperfusion dorsal skin flap model in rats, we show that the hydrogel patch maintains the immediate opening of blood flow channels in transplanted tissue and effective blood perfusion throughout the perioperative period, activating perfusion of the hemodynamic donor site. We demonstrate that the hydrogel patch promotes distal vascularization and long-term functional reconstruction of transplanted tissues by inhibiting inflammatory damage and accelerating blood vessel formation.

NIR-II Imaging and Sandwiched Plasmonic Biosensor for Ultrasensitive Intraoperative Definition of Tumor-Invaded Lymph Nodes.

Radical lymphadenectomy remains the cornerstone of preventing tumor metastasis through the lymphatic system. Current surgical resection of lymph nodes (LNs) based on fluorescence-guided surgery (FGS) suffers from low sensitivity/selectivity with only qualitative information, hampering accurate intraoperative decision-making. Herein, we develop a modularized theranostic system including NIR-II FGS and a sandwiched plasmonic chip (SPC). Intraoperative NIR-II FGS and detection of tumor-positive lymph nodes were performed on the gastric tumor to determine the feasibility of the modularized theranostic system in defining LN metastasis. Under the NIR-II imaging window, the orthotopic tumor and sentinel lymph nodes (SLNs) were successfully excised without ambient light interference in the operating room. Importantly, the SPC biosensor achieved 100% sensitivity and 100% specificity for tumor markers and realized rapid and high-throughput intraoperative SLN detection. We propose the synergetic design of combining the NIR-II FGS and suitable biosensor will substantially improve the efficiency of cancer diagnosis and therapy follow-up.