Chemically Powered Nanomotors with Magnetically Responsive Function for Targeted Delivery of Exosomes.

Janus structure plays a crucial role in achieving chemically driven nanomotors with exceptional motion performance. However, Janus-structured chemically driven nanomotors with magnetic responsiveness are commonly fabricated by sputtering metal films. In the study, a self-assembly technique is employed to asymmetrically modify the surfaces of magnetic silica (SiO2@Fe3O4) nanoparticles with platinum nanoparticles, resulting in the formation of this kind nanomotors. Compared to platinum film, platinum nanoparticles exhibit a larger surface area and a higher catalytic activity. Hence, the nanomotors demonstrate improved diffusion capabilities at a significantly lower concentration (0.05%) of hydrogen peroxide (H2O2). Meanwhile, exosomes have gained attention as a potential tool for the efficient delivery of biological therapeutic drugs due to their biocompatibility. However, the clinical applications of exosomes are limited by their restricted tropism. The previously obtained nanomotors are utilized to deliver exosomes, greatly enhancing its targetability. The drug doxorubicin (DOX) is subsequently encapsulated within exosomes, acting as a representative drug model. Under the conditions of H2O2 concentration at the tumor site, the exosomes exhibited a significantly enhanced rate of entry into the breast cancer cells. The utilization of the nanomotors for exosomes presents a novel approach in the development of hybrid chemically and magnetically responsive nanomotors.

Highly Accurate Profiling of Exosome Phenotypes Using Super-resolution Tricolor Fluorescence Co-localization.

Exosomes contain a wealth of proteomic information, presenting promising biomarkers for the noninvasive early diagnosis of diseases, especially cancer. However, it remains a great challenge to accurately and reliably distinguish exosomes secreted from different types of cell lines. Fluorescence immunoassay is frequently used for exosome detection. Nonspecific adsorption in immunoassays is unavoidable and affects the reliability of assay results. Despite the fact that various methods have been proposed to reduce nonspecific adsorption, a more effective method that can eliminate the influence of nonspecific adsorption is still lacking. Here, we report a more convenient way (named SR-TFC) to remove the artifacts caused by nonspecific adsorption, which combines tricolor fluorescence labeling of target exosomes, tricolor super-resolution imaging, and pixel counting. The pixel counting method (named CFPP) is realized by MATLAB and can eliminate nonspecific binding sites at the single-pixel level, which has never been achieved before and could improve the reliability of detection to the maximum extent. Furthermore, as a proof-of-concept, profiling of exosomal membrane proteins and identification of breast cancer subpopulations are demonstrated. To enable multiplex breast cancer phenotypic analysis, three kinds of specific proteins are labeled to obtain the 3D phenotypic information on various exosomes. Breast cancer subtypes can be accurately identified according to the super-resolution images of some clinically relevant exosomal proteins. Worth mentioning is that, by selecting other biomarkers, classification of other cancers could also be realized using SR-TFC. Hence, the present work holds great potential in clinical cancer diagnosis and precision medicine.

Ultrasensitive Detection of Matrix Metalloproteinase 2 Activity Using a Ratiometric Surface-Enhanced Raman Scattering Nanosensor with a Core-Satellite Structure.

Matrix metalloproteinase 2 (MMP-2) has been considered a promising molecular biomarker for cancer diagnosis due to its related dysregulation. In this work, a core-satellite structure-powered ratiometric surface-enhanced Raman scattering (SERS) nanosensor with high sensitivity and specificity to MMP-2 was developed. The SERS nanosensor was composed of a magnetic bead encapsulated within a 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB)-labeled gold shell as the capture core and a 4-mercaptobenzonitrile (MBN)-encoded silver nanoparticle as the signal satellite, which were connected through a peptide substrate of MMP-2. MMP-2-triggered cleavage of peptides from the core surface resulted in a decrease of the SERS intensity of MBN. Since the SERS intensity of DTNB was used as an internal standard, the reliable and sensitive quantification of MMP-2 activity would be realized by the ratiometric SERS signal, with a limit of detection as low as 2.067 ng/mL and a dynamic range from 5 to 100 ng/mL. Importantly, the nanosensor enabled a precise determination of MMP-2 activity in tumor cell secretions, which may provide an avenue for early diagnosis and classification of malignant tumors.

A SERS-assisted 3D organotypic microfluidic chip for in-situ visualization and monitoring breast cancer extravasation process.

Extravasation, as one of the key steps in cancer metastasis, refers to the process where tumor cells escape the bloodstream by crossing the vascular endothelium and invade the targeted tissue, which accounts for the low five-year survival rate of cancer patients. Understanding the mechanism of cancer metastasis and inhibiting extravasation are crucial to improve patient prognosis. Here, a 3D organotypic microfluidic chip combined with SERS-based protein imprinted nanomaterials (SPINs) was proposed to study the extravasation process in vitro. The chip consists of a collagen gel channel and a vascular channel where human vein endothelial cells (HUVECs) and breast cancer cells are injected sequentially to induce extravasation. By comparing two subtypes of breast cancer cells (MCF-7 and MDA-MB-231), we successfully observed the difference in extravasation capabilities between two kinds of cells through fluorescence imaging. Meanwhile, thanks to the high specificity of molecular imprinting technology and the high sensitivity of surface enhanced Raman scattering (SERS), SPINs were utilized to analyze the concentration of several cancer secretions (interleukin-6 and interleukin-8) in complex biological fluid in real-time. Further, our model showed that downregulation of secretions by therapeutic drugs can inhibit the extravasation of breast cancers. This microfluidic model may pave the way for the fundamental research of the cancer metastasis and evaluating the therapeutic efficacy of potential drugs.

Long-lived SERS Matrix for Real-Time Biochemical Detection Using "Frozen" Transition State.

For the long-time tracking of biological events, maintaining the bioactivity of the analytes during the detection process is essential. Here, we show a versatile surface-enhanced Raman Scattering (SERS) platform, termed a superwettable-omniphobic lubricous porous SERS (SOLP-SERS) substrate. The SOLP-SERS substrate could generate a three-dimensional liquid "hotspots" matrix with an ultra-long lifetime (tens of days) by confining tiny amounts of liquids within the gaps between nanoparticles. Then, the analytes are trapped in the uniform liquid "hotspots", whose bioactivity can be well maintained over a long period of time during SERS detection. Limits of detection down to femtomolar levels were achieved for various molecules. More importantly, SERS signals were uniform within the substrate and remained stable for more than 30 days. As a proof-of-concept experiment, the dynamic detection of the polymerization of Aß peptides into amyloids was monitored by the SOLP-SERS substrate within 48 h. Moreover, the exosomes secreted by breast cancer cells, an important biomarker of cancer, were also measured. These results demonstrate that the SOLP-SERS platform will provide new insights into the development of real-time biochemical sensors with ultrahigh sensitivity.

A 3D-printed SERS bionic taster for dynamic tumor metabolites detection.

The variation of tumor-associated metabolites in extracellular microenvironment timely reflects the development, the progression and the treatment of cancers. Conventional methods for metabolite detection lack the efficiency to grasp the dynamic metabolic alterations. Herein, we developed a SERS bionic taster which enabled real-time analysis of extracellular metabolites. The instant information of cell metabolism was provided by the responsive Raman reporters, which experienced SERS spectral changes upon metabolite activation. Such a SERS sensor was integrated into a 3D-printed fixture which fits the commercial-standard cell culture dishes, allowing in-situ acquisition of the vibrational spectrum. The SERS taster can not only accomplish simultaneous and quantitative analysis of multiple tumor-associated metabolites, but also fulfill the dynamic monitoring of cellular metabolic reprogramming, which is expected to become a promising tool for investigating cancer biology and therapeutics.

Nanoscale imaging of tumor cell exosomes by expansion single molecule localization microscopy (ExSMLM).

Tumor cell exosomes play a very important role in the process of tumor cell proliferation and metastasis. However, due to the nanoscale size and high heterogeneity of exosomes, in-depth understanding of their appearance and biological characteristics is still lacking. Expansion microscopy (ExM) is a method that embeds biological samples in a swellable gel to physically magnify the samples to improve the imaging resolution. Before the emergence of ExM, scientists had invented several super-resolution imaging techniques that could break the diffraction limit. Among them, single molecule localization microscopy (SMLM) usually has the best spatial resolution (20-50 nm). However, considering the small size of exosomes (30-150 nm), the resolution of SMLM is still not high enough for detailed imaging of exosomes. Hence, we propose a tumor cell exosomes imaging method that combines ExM and SMLM (i.e. Expansion SMLM, denoted as ExSMLM), which can realize the expansion and super-resolution imaging of tumor cell exosomes. In this technique, immunofluorescence was first performed to fluorescently label the protein markers on the exosomes, then the exosomes were polymerized into a swellable polyelectrolyte gel. The electrolytic nature of the gel made the fluorescently labeled exosomes undergo isotropic linear physical expansion. The expansion factor obtained in the experiment was about 4.6. Finally, SMLM imaging of the expanded exosomes was performed. Owing to the improved resolution of ExSMLM, nanoscale substructures of closely packed proteins were observed on single exosomes, which has never been achieved before. With such a high resolution, ExSMLM would have a great potential in detailed investigation of exosomes and exosome-related biological processes.

Red Blood Cell Membrane Camouflaged Mesoporous Silica Nanorods as Nanocarriers for Synergistic Chemo-Photothermal Therapy.

In recent years, nanoparticles camouflaged by red blood cell membrane (RBCM) have become a potential nano-drug delivery platform due to their good biocompatibility and immune evasion capability. Here, a multifunctional drug nanocarrier based on RBCM camouflaged mesoporous silica nanorods (MSNR) is presented, which can be used in pH and near-infrared (NIR) light triggered synergistic chemo-photothermal killing of cancer cells. To fabricate such a nanocarrier, MSNR and RBCM were prepared by the sol-gel method and modified hypotonic lysis method, respectively. Drugs were loaded into the pores of MSNR. Finally, RBCM was coated on the surface of MSNR by extrusion through a polycarbonate membrane. The advantages of the nanocarrier include: 1) MSNR can induce more cellular uptake than sphere shaped mesoporous silica nanoparticles. 2) The RBCM can reduce drug leakage and prevent clearance of the nanocarriers by macrophages. 3) By simultaneous loading doxorubicin (DOX) and indocyanine green (ICG), pH and NIR triggered synergistic chemo-photothermal therapy can be realized. In the experiment, we studied the drug releasing and cellular uptake of the nanocarriers in a breast cancer cell line (SKBR3 cells), in which a sufficient killing effect was observed. Such a multifunctional drug nanocarrier holds a broad application prospect in cancer treatment.

Metal nanoprobe-decorated all-inorganic perovskite nanocrystal-based fluorescence-linked immunosorbent assay for the detection of tumor-derived exosomes.

All-inorganic perovskite nanocrystals (CsPbX3 NCs, X = Cl, Br, I) are promising fluorescence materials for biological detection due to their excellent optical properties. However, there is still a challenge to obtain stable CsPbX3 NCs with more biofunctions. Here, we proposed a distinct strategy by absorbing the functionalized metal nanoprobes onto the phospholipid encapsulated CsPbX3 NCs to achieve CsPbX3-metal hybrids as probes for the detection of tumor-derived exosomes. Here, the metal nanoprobes have two functions: first, it endows phospholipid encapsulated CsPbX3 NCs with recognition ability; second, it avoids the fluorescence quenching of CsPbX3 NCs during the biological modification process by using metal nanoparticles as a bridge to connect with CsPbX3 NCs and various biomolecules. The obtained CPXD-AD exhibited a bright fluorescence signal, narrow full width at half-maximum (FWHM), and high specificity. Under optimal conditions, the CPXD-AD-based fluorescence-linked immunosorbent assay (FLISA) was successfully established and used for both qualitative and quantitative detection of tumor-derived exosomes.

Fluorescence Super-Resolution Imaging Chip for Gene Silencing Exosomes.

Tumor cell-derived extracellular vesicles and their cargo of bioactive substances have gradually been recognized as novel biomarkers for cancer diagnosis. Meanwhile, the PD-L1 (Programmed Death-Ligand 1) protein, as an immune checkpoint molecule, is highly expressed on certain tumor cells and holds significant potential in immune therapy. In comparison to PD-L1 monoclonal antibodies, the inhibitory effect of PD-L1 siRNA (small interfering RNA) is more advantageous. In this article, we introduced a microfluidic chip integrating cell cultivation and exosome detection modules, which were intended for the investigation of the gene silencing effect of PD-L1 siRNA. Basically, cells were first cultured with PD-L1 siRNA in the chip. Then, the secreted exosomes were detected via super-resolution imaging, to validate the inhibitory effect of siRNA on PD-L1 expression. To be specific, a "sandwich" immunological structure was employed to detect exosomes secreted from HeLa cells. Immunofluorescence staining and DNA-PAINT (DNA Point Accumulation for Imaging in Nanoscale Topography) techniques were utilized to quantitatively analyze the PD-L1 proteins on HeLa exosomes, which enabled precise structural and content analysis of the exosomes. Compared with other existing PD-L1 detection methods, the advantages of our work include, first, the integration of microfluidic chips greatly simplifying the cell culture, gene silencing, and PD-L1 detection procedures. Second, the utilization of DNA-PAINT can provide an ultra-high spatial resolution, which is beneficial for exosomes due to their small sizes. Third, qPAINT could allow quantitative detection of PD-L1 with better precision. Hence, the combination of the microfluidic chip with DNA-PAINT could provide a more powerful integrated platform for the study of PD-L1-related tumor immunotherapy.

In Situ Super-Resolution Imaging of Telomeres with DNA-PAINT.

Telomeres are located at the ends of chromosomes and play an important role in maintaining the integrity of chromosomes and controlling the cycle of cell division. Studies have shown that abnormal telomere length may lead to the occurrence of some diseases. Therefore, accurate measurement of telomere length will be helpful for the prediction and diagnosis of related diseases. DNA point accumulation for imaging in nanoscale topography (PAINT) is an optical super-resolution technology that relies on the instantaneous binding of the fluorescent DNA imaging strand to the target epitope. Here, we present the first demonstration of DNA-PAINT-based in situ super-resolution imaging of telomeres as well as centromeres. For DNA-PAINT imaging, Cy5-labeled telomere DNA (5'-Cy5-TTTTTCCCTAACCCTAA-3') and Cy3-labeled centromere DNA (5'-Cy3-TTTTTAGCTTCTGTCTAGTTT-3') are utilized as the imager strands. Through an improved permeabilization strategy that we proposed, the imager strands can bind with intracellular telomeres and centromeres with high specificity, realizing super-resolution imaging of telomeres and centromeres. To check the applicability of DNA-PAINT in evaluating telomere length, we conducted an experiment using azidothymidine (AZT)-treated tumor cells as the imaging target. The DNA-PAINT imaging results clearly revealed the telomerase inhibition effect of AZT. Compared with single-molecule localization microscopy (SMLM) with peptide nucleic acid (PNA)-based fluorescence in situ hybridization (FISH), our method has the advantages of low cost, low toxicity, and simple equipment. Such a DNA-PAINT-based imaging strategy holds great potential in measuring telomere length with high accuracy, which would play an important role in the study of telomere-related diseases such as cancer.

A Programmable Plasmonic Gas Microsystem for Detecting Arbitrarily Combinated Volatile Organic Compounds (VOCs) with Ultrahigh Resolution.

For gas sensors, the ultrasensitive and highly selective detection of multiple components is of great significance in a wide range of applications extending from environment to healthcare, which is still a long-term challenge due to the single sensing mechanism of most sensors. Here, we combine the advantages of microfluidic chips and surface-enhanced Raman spectroscopy (SERS) spectra to fabricate a smart single-chip for simultaneously detecting an arbitrary combination of VOCs that incorporates different detection units, working on either a physisorption or chemisorption mechanism. Full integration of microfluidic and multiplex nanostructure components on one chip permits programmable design for sensing multifarious volatile compounds, and enables on-chip signal amplifications with increased reproducibility. As a proof-of-principle experiment, we demonstrate the simultaneous identification of 9 different gases that belong to aromatic compounds, aldehydes, ketones, or sulfides in one mixture, with high sensitivity (ppb level), high selectivity, and high robustness (error ∼8%). We further evaluated the application of our universal gas sensor in two scenarios including indoor air pollution monitoring and exhaled breath-based disease diagnosis. We expect that our design will improve the various practical applications of gas sensors.

Quantitative analysis of multiple breast cancer biomarkers using DNA-PAINT.

Immunotherapy has become an efficient treatment method of breast cancer. Detection of proteins such as PD-L1 and CTLA-4, which are important immune checkpoint molecules, is attracting more and more attention as they play key roles in immunotherapy. Here, by combining the high resolution of DNA-PAINT (DNA points accumulation for imaging in nanoscale topography) with the qPAINT quantitative analysis method, accurate spatial localization and absolute quantification of PD-L1 and CTLA-4 on the membrane of breast cancer cells could be achieved. Meanwhile, exchange-PAINT was also conducted to count three other biomarkers (EpCAM, EGFR, and HER2). Simultaneous analysis of these biomarkers can greatly facilitate the differentiation of different kinds of breast cancer. Such a simple quantitative analysis method holds great potential in diagnosis and immunotherapy of cancers.

Triple-color fluorescence co-localization of PD-L1-overexpressing cancer exosomes.

Programed cell death ligand 1 (PD-L1) is a protein biomarker overexpressed on exosomes derived from tumor cells. It plays an important role in tumor diagnosis, screening, evaluation of therapeutic efficacy, and prognosis. In this study, a facile method is presented to detect PD-L1-overexpressing cancer exosomes with high specificity and sensitivity. First, gold nanospheres (GNSs) were attached to the bottom of an eight-well chambered slide by electrostatic adsorption, forming the detection substrate. Then, Cy5-labeled CD63 aptamers (i.e., the capture probes) were modified on the GNSs by Au-S bond. After adding samples containing target exosomes which were stained by membrane dyes DiI in advance, FAM-labeled PD-L1 aptamers (i.e., the immunoprobes) were added to recognize PD-L1 on the target exosomes. By triple-color fluorescence co-localization (TFC) of the Cy5, DiI, and FAM channels, highly sensitive and reliable detection of the PD-L1-overexpressing exosomes was achieved in the concentration range 7.78 × 101 to 7.78 × 104 particles/mL with a detection limit down to 6 particles/mL. The advantages of the proposed detection method include the following; first, the detection substrate is easy to prepare and convenient to clean. Second, the TFC strategy can completely exclude nonspecific reaction sites and thus significantly improves the accuracy. Such a facile and reliable detection method holds a great potential in exosome-based cancer theranostics. In this paper, we proposed a triple-color fluorescence co-localization (TFC) strategy to significantly improve the reliability of exosome detection and the detection substrate is easy to prepare and convenient to clean. In addition, the LOD is down to 6 particles/mL, which is quite low compared with other detection methods.

A Ti2N MXene-based nanosystem with ultrahigh drug loading for dual-strategy synergistic oncotherapy.

The exploration of MXenes, especially nitride MXenes, in the field of theranostic nanomedicine is still in its infancy. Here, towards synergistic chemo-photothermal oncotherapy, we demonstrate the first kind of 2D titanium nitride (Ti2N) MXene-based nanosystem (Ti2N@oSi) for dual-strategy synergistic oncotherapy. The unique structure of Ti2N nanosheets endows the drug carriers with an ultrahigh loading capacity of 796.3% and an excellent NIR photothermal conversion efficiency of 41.6% for chemo-photothermal therapy. After being coated with a biodegradable organosilica shell, the Ti2N@oSi nanocarriers show excellent characteristics of tumor targeting, pH/glutathione/photothermal-responsive drug release and dual-drug combination chemotherapy. Both in vitro and in vivo therapeutic evaluations demonstrate the pronounced tumor growth inhibition effect and superior biocompatibility of Ti2N@oSi nanocarriers. The excellent drug loading ability, photothermal conversion ability and surface modifiability of Ti2N open up new opportunities for tumor microenvironment-targeted synergistic oncotherapy. This work is supposed to broaden the application of MXenes in nanomedicine and, particularly, provide the first sight to the biomedical application of nitride MXenes.

Telomerase detection using a DNA-PAINT strategy.

Telomerase plays an important role in maintaining the length of telomere during cell division and is recognized as a new kind of biomarkers for cancer diagnosis. In this work, we present a brand new telomerase detection strategy based on a DNA points accumulation for imaging in nanoscale topography (DNA-PAINT) like strategy. With an extraordinary spatial resolution (∼10 nm), the DNA-PAINT based strategy offers several advantages. First, it avoids complicated polymerase chain reaction and electrophoresis procedures. Second, it enables super resolution imaging of the reaction products with a high signal-to-noise ratio and facilitates the location of telomeric elongation sites on the single particle level, which results in a high sensitivity. Third, the detection scheme of the DNA-PAINT strategy allows directin situvisualization of the telomeric elongation process, which has never been achieved before. All these advantages make the DNA-PAINT telomerase detection strategy significant for dynamic investigation of telomerase related physiological processes as well as cancer diagnosis.

Ti3C2Tx MXene-Loaded 3D Substrate toward On-Chip Multi-Gas Sensing with Surface-Enhanced Raman Spectroscopy (SERS) Barcode Readout.

Gas sensors lie at the heart of various fields ranging from medical to environmental sciences, and the demand of gas sensors is instantly expanding. However, in the face of complex gas samples, how to maintain high sensitivity while performing multiplex detection still puzzles the researchers. Here, by introducing Ti3C2Tx MXene into a microfluidic gas sensor with a three-dimensional (3D) transferable SERS substrate, a powerful gas sensor having both multiplex detecting ability and high sensitivity is demonstrated. The employ of MXene endows the sensor with a universal high adsorption efficiency for various gases while the generation of in situ gas vortices in the sophisticated nanomicro structure extends the molecule residence time in SERS-active area, both leading to the increased sensitivity. In the proof-of-concept experiment, a limit of detection (LOD) of 10-50 ppb was achieved for three typical volatile organic compounds (VOCs) according to the intrinsic SERS signals of gas molecules. Besides, the well-designed periodic 3D structure solves the general repeatability problem of SERS substrates. In addition, the detailed composition of gas mixture was revealed using classic least-square analysis (CLS) with an average accuracy of 90.6%. Further, a chromatic barcode was developed based on the results of CLS to read out the complex composition of samples visually.

Eliminating nonspecific binding sites for highly reliable immunoassay via super-resolution multicolor fluorescence colocalization.

Non-specific adsorption in immunoassays has always been a major problem that affects the reliability of assay results. Despite the emergence of various methods that can reduce nonspecific adsorption, a universal and effective method to reduce the influence of nonspecific adsorption is still lacking. Hence, we propose here an optical super-resolution imaging based immunoassay strategy, named super-resolution multicolor fluorescence colocalization (SR-MFC), which can generate a low false-positive rate. Taking advantages of the high spatial resolution of single-molecule localization microscopy (SMLM), SR-MFC can directly visualize the assay results and thus effectively exclude the nonspecific binding sites. In other words, even if nonspecific interactions do happen, SR-MFC ensures that the nonspecific reaction sites are visualized and abandoned, which has never been achieved before. To verify its practicability, exosomes, which are important cancer biomarkers, were used as model targets and detected using SR-MFC. Compared with common immunofluorescence assay, the accuracy and reliability of the detection results are greatly improved. The detection limit of exosomes was 38 particles per µL. More importantly, the SR-MFC method can also be generalized for the detection of other biomarkers (e.g. proteins, DNAs, etc.), which is a significant and promising new strategy for immunoassay based diagnosis.

Simultaneous detection of multiple exosomal microRNAs for exosome screening based on rolling circle amplification.

Exosomal microRNAs (miRNAs) have attracted great attention as predictive and prognostic biomarkers of cancer. Profiling of miRNAs plays a key role in the effective diagnosis of cancers. However, simultaneous quantification of multiple miRNAs is challenging due to their homology and low abundance especially in exosomes. Here, we developed a sensitive detection method for multiple exosomal miRNAs with the help of rolling circle amplification (RCA). In contrast of the traditional ways, this method takes the advantages of both the multiplex sensing ability and the simplicity of RCA. Specifically, multiple exosomal miRNAs from different cell lines were replicated simultaneously through RCA and detected using designed molecular beacons (MBs). miRNA-21, miRNA-122 and miRNA-155 were chosen as the targets, which are overexpressed in cancers. Normalized fluorescence intensities of MB were used to imply the relative concentrations of these miRNAs. The obtained relative miRNAs expression levels could be used to distinguish the breast cancer exosome from normal one. If the varieties of the detected exosomal miRNAs are abundant enough, the concentration ratios of miRNAs could basically indicate the corresponding exosome and exosome screening could be realized. Such exosomal miRNA profiling and exosome screening can assist cancer diagnosis, which is promising in clinical application.

SERS-fluorescence-superresolution triple-mode nanoprobe based on surface enhanced Raman scattering and surface enhanced fluorescence.

Multifunctional nanoprobes play important roles in cell imaging and sensing. Here, we present a novel optical nanoprobe based on surface enhanced Raman scattering (SERS) and surface enhanced fluorescence (SEF), which can realize the SERS-fluorescence and superresolution triple-mode imaging of cancer cells. Compared with other previously reported multifunctional nanoprobes, the proposed nanoprobe holds two exquisite properties. The first one is that, in addition to normal SERS and fluorescence imaging, the nanoprobe can also be used for single molecule localization microscopy (SMLM) imaging, which helps compensate for the diffraction limited spatial resolution of normal SERS and fluorescence imaging. The second one is that, other than simple fluorescence, SEF is used in the nanoprobe to produce a stronger signal for fluorescence imaging and, more importantly, better photo-switching for SMLM imaging. In the experiment, we optimized the structure of the nanoprobe to obtain the best SEF effect. With the optimal structure, the triple-mode imaging of a breast cancer cell line (SKBR3) is realized. Since such triple-mode imaging of cancer cells has never been achieved before, we believe that the presented nanoprobe holds great potential for cancer cell targeting or the investigation of cell-nanomaterial interactions.