Cancer screening through surface-enhanced Raman spectroscopy fingerprinting analysis of urinary metabolites using surface-carbonized silver nanowires on a filter membrane.

BACKGROUND: Label-free surface-enhanced Raman spectroscopy (SERS)-based metabolic profiling has great potential for early cancer diagnosis, but further advancements in analytical methods and clinical evidence studies are required for clinical applications. To improve the cancer diagnostic accuracy of label-free SERS spectral analysis of complex biological fluids, it is necessary to obtain specifically enhanced SERS signals of cancer-related metabolites present at low concentrations. RESULTS: This study presents a novel 3D SERS sensor, comprising a surface-carbonized silver nanowire (AgNW)-stacked filter membrane, alongside an optimized urine/methanol/chloroform extraction technique, which specifically changes the molecular adsorption and orientation of aromatic metabolites onto SERS substrates. By analyzing the pretreated urine samples on the surface-carbonized AgNW 3D SERS sensor, distinct and highly enhanced SERS peaks derived from semi-polar aromatic metabolites were observed for pancreatic cancer and prostate cancer samples compared with normal controls. Urine metabolite analysis using SERS fingerprinting successfully differentiated pancreatic cancer and prostate cancer groups from normal control group: normal control (n = 56), pancreatic cancer (n = 40), and prostate cancer (n = 39). SIGNIFICANCE AND NOVELTY: We confirmed the clinical feasibility of performing fingerprint analysis of urinary metabolites based on the surface-carbonized AgNW 3D SERS sensor and methanol/chloroform extraction for noninvasive cancer screening. This technology holds potential for large-scale screening owing to its high accuracy, and cost effective, simple and rapid detection method.

Metal-enhanced fluorescence biosensor integrated in capillary flow-driven microfluidic cartridge for highly sensitive immunoassays.

Point-of-care testing (POCT) for low-concentration protein biomarkers remains challenging due to limitations in biosensor sensitivity and platform integration. This study addresses this gap by presenting a novel approach that integrates a metal-enhanced fluorescence (MEF) biosensor within a capillary flow-driven microfluidic cartridge (CFMC) for the ultrasensitive detection of the Parkinson's disease biomarker, aminoacyl-tRNA synthetase complex interacting multi-functional protein 2 (AIMP-2). Crucial point to this approach is the orientation-controlled immobilization of capture antibody on a nanodimple-structured MEF substrate within the CFMC. This strategy significantly enhances fluorescence signals without quenching, enabling accurate quantification of low-concentration AIMP-2 using a simple digital fluorescence microscope with a light-emitting diode excitation source and a digital camera. The resulting platform exhibits exceptional sensitivity, achieving a limit of detection in the pg/mL range for AIMP-2 in human serum. Additionally, the CFMC design incorporates a capillary-driven passive sample transport mechanism, eliminating the need for external pumps and further simplifying the detection process. Overall, this work demonstrates the successful integration of MEF biosensing with capillary microfluidics for point-of-care applications.

Micromolding-Assisted Production of SERS-Active Microcylinders for Size- and Charge-Selective Molecular Detection.

Surface-enhanced Raman scattering (SERS) is an effective technique for amplifying the Raman signal of molecules by using metal nanostructures. However, these metal surfaces are susceptible to contamination by undesirable adhesives in complex mixtures, typically necessitating a time-consuming and costly sample pretreatment. In order to circumvent this, metal nanoparticles have been uniformly embedded within microgels by using microfluidics. In this work, we introduce a simple, scalable micromolding method for creating SERS-active cylindrical microgels designed to eliminate the need for pretreatment. These microcylinders are created through the simultaneous photoreduction and photo-cross-linking of precursor solutions. These solutions are optimized for consistent, high-intensity Raman signals as well as molecular size and charge selectivity. A sequential micromolding method is employed to design dual-compartment microcylinders, offering additional functionalities such as optical encoding, magnetoresponsiveness, and dual-charge selectivity. These SERS-active microcylinders provide robust Raman signals of small molecules, even in the presence of adhesive proteins, without compromising sensitivity. To demonstrate this capability, we directly detect pyocyanin in saliva and tartrazine in whole milk without any need for sample pretreatment.

Label-free detection and discrimination of respiratory pathogens based on electrochemical synthesis of biomaterials-mediated plasmonic composites and machine learning analysis.

Seasonal outbreaks of respiratory viral infections remain a global concern, with increasing morbidity and mortality rates recorded annually. Timely and false responses contribute to the widespread of respiratory pathogenic diseases owing to similar symptoms at an early stage and subclinical infection. The prevention of emerging novel viruses and variants is also a big challenge. Reliable point-of-care diagnostic assays for early infection diagnosis play a critical role in the response to threats of epidemics or pandemics. We developed a facile method for specifically identifying different viruses based on surface-enhanced Raman spectroscopy (SERS) with pathogen-mediated composite materials on Au nanodimple electrodes and machine learning (ML) analyses. Virus particles were trapped in three-dimensional plasmonic concave spaces of the electrode via electrokinetic preconcentration, and Au films were simultaneously electrodeposited, leading to the acquisition of intense and in-situ SERS signals from the Au-virus composites for ultrasensitive SERS detection. The method was useful for rapid detection analysis (<15 min), and the ML analysis for specific identification of eight virus species, including human influenza A viruses (i.e., H1N1 and H3N2 strains), human rhinovirus, and human coronavirus, was conducted. The highly accurate classification was achieved using the principal component analysis-support vector machine (98.9%) and convolutional neural network (93.5%) models. This ML-associated SERS technique demonstrated high feasibility for direct multiplex detection of different virus species for on-site applications.

Dual-Enhanced Plasmonic Biosensing for Point-of-Care Sepsis Detection.

Rapid, sensitive, simultaneous quantification of multiple biomarkers in point-of-care (POC) settings could improve the diagnosis and management of sepsis, a common, potentially life-threatening condition. Compared to high-end commercial analytical systems, POC systems are often limited by low sensitivity, limited multiplexing capability, or low throughput. Here, we report an ultrasensitive, multiplexed plasmonic sensing technology integrating chemifluorescence signal enhancement with plasmon-enhanced fluorescence detection. Using a portable imaging system, the dual chemical and plasmonic amplification enabled rapid analysis of multiple cytokine biomarkers in 1 h with sub-pg/mL sensitivities. Furthermore, we also developed a plasmonic sensing chip based on nanoparticle-spiked gold nanodimple structures fabricated by wafer-scale batch processes. We used the system to detect six cytokines directly from clinical plasma samples (n = 20) and showed 100% accuracy for sepsis detection. The described technology could be employed in rapid, ultrasensitive, multiplexed plasmonic sensing in POC settings for myriad clinical conditions.

3D plasmonic coral nanoarchitecture paper for label-free human urine sensing and deep learning-assisted cancer screening.

Practical human biofluid sensing requires a sensor device to differentiate patients from the normal group with high sensitivity and specificity. Label-free molecular identification from human biofluids allows direct classification of abnormal samples, providing insights for disease diagnosis and finding of new biomarkers. Here, we introduce a label-free surface-enhanced Raman scattering sensor based on a three-dimensional plasmonic coral nanoarchitecture (3D-PCN), which has strong electromagnetic field enhancement through multiple hot spots. The 3D-PCN was synthesized on a paper substrate via direct one-step gold reduction, forming a coral-like nanoarchitecture with high absorption property for biofluids. This was fabricated as a urine test strip and then integrated with a handheld Raman system to develop an on-site urine diagnostic platform. The developed platform successfully classified the human prostate and pancreatic cancer urines in a label-free method supported by two types of deep learning networks, with high clinical sensitivity and specificity. Our technology has the potential to be utilized not only for urinary cancer diagnosis but also for various human biofluid sensing systems as a future point-of-care testing platform.

In-situ fabrication of 3D interior hotspots templated with a protein@Au core-shell structure for label-free and on-site SERS detection of viral diseases.

Nanoscale plasmonic hotspots play a critical role in the enhancement of molecular Raman signals, enabling the sensitive and reliable trace analysis of biomedical molecules via surface-enhanced Raman spectroscopy (SERS). However, effective and label-free SERS diagnoses in practical fields remain challenging because of clinical samples' random adsorption and size mismatch with the nanoscale hotspots. Herein, we suggest a novel SERS strategy for interior hotspots templated with protein@Au core-shell nanostructures prepared via electrochemical one-pot Au deposition. The cytochrome c and lysates of SARS-CoV-2 (SLs) embedded in the interior hotspots were successfully functionalized to confine the electric fields and generate their optical fingerprint signals, respectively. Highly linear quantitative sensitivity was observed with the limit-of-detection value of 10-1 PFU/mL. The feasibility of detecting the targets in a bodily fluidic environment was also confirmed using the proposed templates with SLs in human saliva and nasopharyngeal swabs. These interior hotspots templated with the target analytes are highly desirable for early and on-site SERS diagnoses of infectious diseases without any labeling processes.

Interior Hotspot Engineering in Ag-Au Bimetallic Nanocomposites by In Situ Galvanic Replacement Reaction for Rapid and Sensitive Surface-Enhanced Raman Spectroscopy Detection.

Engineering of interior hotspots provides a paradigm shift from traditional surface-enhanced Raman spectroscopy (SERS), in which the detection sensitivity depends on the positioning of adsorbed molecules. In the present work, we developed an Ag-Au bimetallic nanocomposite (SGBMNC) SERS platform with interior hotspots through facile chemical syntheses. Ag nanoparticles replaced by Au via the galvanic replacement reaction (GRR) provided hotspot regions inside the SGBMNC that remarkably enhanced the plasmonic activity compared to the conventional SERS platforms without the internal hotspots. The diffusion of analytes into the proposed interior hotspots during the GRR process enabled sensitive detections within 10 s. The SERS behaviors of the SGBMNC platform were investigated using methylene blue (MB) as a Raman probe dye. A quantitative study revealed excellent detection performance, with a limit of detection (LOD) of 42 pM for MB dye and a highly linear correlation between peak intensity and concentration (R2 ≥ 0.91). The SGBMNC platform also enabled the detection of toxic benzyl butyl phthalate with a sufficient LOD of 0.09 ppb (i.e., 280 pM). Therefore, we believe that the proposed methodology can be used for SERS assays of hazardous materials in practical fields.

Organometallic hotspot engineering for ultrasensitive EC-SERS detection of pathogenic bacteria-derived DNAs.

The sensitivity and limit-of-detection (LOD) of the traditional surface-enhanced Raman spectroscopy (SERS) platform suffer from the requirement of precise positioning of small analytes, including DNAs and bacteria, into narrow hotspots. In this study, a novel SERS sensor was developed using electrochemical deposition onto metal nanopillars (ECOMPs) combined with complementary DNAs (cDNAs) for the detection of pathogenic bacteria. Applying a redox potential to AuCl4- ions actively engineered the organometallic hotspots based on the cDNAs in a short time (<10 min) and simultaneously produced SERS signals. Because of the influence of potential-driven morphological properties on the SERS efficiency in the cDNA domains and the resonant coupling of internal fields with the fields confined between adjacent ECOMPs-cDNAs, the optimum growth time was determined to be 5 min. The EC-SERS detection and discrimination of Enterococcus faecium and Staphylococcus aureus were successfully carried out because of the DNA complementarity. Compared with plasmonic metal nanopillars (MPs)-cDNAs, the enhancement factor of the ECOMPs-cDNAs was estimated to be ∼2.0 × 103. A quantitative investigation revealed that a highly linear progression in the target DNA concentration range (0.05-100 nM) and a LOD of ∼0.035 nM were achieved. The specificity of the ECOMPs-cDNAs was validated by cross-hybridization. The platform was also used to assay human whole blood containing 0.1 nM bacterial DNAs. The proposed strategy provides the potential for highly sensitive SERS-based multiplex DNA detection in clinical diagnostics.

Surface-enhanced Raman scattering-based immunoassay for severe acute respiratory syndrome coronavirus 2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected humans worldwide for over a year now. Although various tests have been developed for the detection of SARS-CoV-2, advanced sensing methods are required for the diagnosis, screening, and surveillance of coronavirus disease 2019 (COVID-19). Here, we report a surface-enhanced Raman scattering (SERS)-based immunoassay involving an antibody pair, SERS-active hollow Au nanoparticles (NPs), and magnetic beads for the detection of SARS-CoV-2. The selected antibody pair against the SARS-CoV-2 antigen, along with the magnetic beads, facilitates the accurate direct detection of the virus. The hollow Au NPs exhibit strong, reproducible SERS signals, allowing sensitive quantitative detection of SARS-CoV-2. This assay had detection limits of 2.56 fg/mL for the SARS-CoV-2 antigen and 3.4 plaque-forming units/mL for the SARS-CoV-2 lysates. Furthermore, it facilitated the identification of SARS-CoV-2 in human nasopharyngeal aspirates and diagnosis of COVID-19 within 30 min using a portable Raman device. Thus, this assay can be potentially used for the diagnosis and prevention of COVID-19.

Hydrogel-Assisted 3D Volumetric Hotspot for Sensitive Detection by Surface-Enhanced Raman Spectroscopy.

Effective hotspot engineering with facile and cost-effective fabrication procedures is critical for the practical application of surface-enhanced Raman spectroscopy (SERS). We propose a SERS substrate composed of a metal film over polyimide nanopillars (MFPNs) with three-dimensional (3D) volumetric hotspots for this purpose. The 3D MFPNs were fabricated through a two-step process of maskless plasma etching and hydrogel encapsulation. The probe molecules dispersed in solution were highly concentrated in the 3D hydrogel networks, which provided a further enhancement of the SERS signals. SERS performance parameters such as the SERS enhancement factor, limit-of-detection, and signal reproducibility were investigated with Cyanine5 (Cy5) acid Raman dye solutions and were compared with those of hydrogel-free MFPNs with two-dimensional hotspots. The hydrogel-coated MFPNs enabled the reliable detection of Cy5 acid, even when the Cy5 concentration was as low as 100 pM. We believe that the 3D volumetric hotspots created by introducing a hydrogel layer onto plasmonic nanostructures demonstrate excellent potential for the sensitive and reproducible detection of toxic and hazardous molecules.

SERS-PCR assays of SARS-CoV-2 target genes using Au nanoparticles-internalized Au nanodimple substrates.

The reverse transcription-polymerase chain reaction (RT-PCR) method has been adopted worldwide to diagnose severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Although this method has good sensitivity and specificity, there is a need to develop a more rapid diagnostic technology, given the virus's rapid spread. However, the RT-PCR method takes a long time to diagnose SARS-CoV-2 because of the required thermocycling steps. Therefore, we developed a surface-enhanced Raman scattering (SERS)-PCR detection method using an AuNP-internalized Au nanodimple substrate (AuNDS) to shorten the diagnosis time by reducing the number of thermocycling steps needed to amplify the DNA. For the representative target markers, namely, the envelope protein (E) and RNA-dependent RNA polymerase (RdRp) genes of SARS-CoV-2, 25 RT-PCR thermocycles are required to reach a detectable threshold value, while 15 cycles are needed for magnetic bead-based SERS-PCR when the initial DNA concentration was 1.00× 105 copies/µL. However, only 8 cycles are needed for the AuNDS-based SERS-PCR. The corresponding detectable target DNA concentrations were 3.36 × 1012, 3.28 × 109, and 2.56 × 107 copies/µL, respectively. Therefore, AuNDS-based SERS-PCR is seen as being a new molecular diagnostic platform that can shorten the time required for the thermocycling steps relative to the conventional RT-PCR.

SERS-based serodiagnosis of acute febrile diseases using plasmonic nanopopcorn microarray platforms.

We report a surface-enhanced Raman scattering (SERS)-based immunoassay platform for the rapid diagnosis of scrub typhus and murine typhus, which are the most common acute febrile diseases in South Korea. A microarray device, composed of multiple gold nanopopcorn substrates capable of detecting ultra-sensitive biomarkers, was used as a multiplex SERS-based assay platform. Sequentially diluted titers of Orientia tsutsugamushi and Rickettsia typhi specific human IgG/IgM antibodies, which are biomarkers of two typhus diseases, were analyzed by Raman spectroscopy, and the peak intensity was plotted against the different titer concentration range (0-2048 and 0-1024 for O. tsutsugamushi IgG/IgM and 0-8192 and 0-256 for R. typhi IgG/IgM) to generate calibration curves. The sensitivities and limits of detection (LODs) determined for four different IgG/IgM antibodies were significantly lower than those for the ELISA method. The LODs of titer concentrations for O. tsutsugamushi IgG/IgM and R. typhi IgG/IgM are determined to be 20.4, 7.03, 16.8 and 12.5, respectively. The LOD values were all lower than the cut-off values (256, 16, 128, and 64) used for clinical diagnosis, which means that this assay platform can diagnose two typhus diseases with high sensitivity. When the microarray sensors are combined with portable Raman spectrophotometers, which are commercially available worldwide, it is also possible to directly diagnose a patient in the field without sending their blood sample to a hospital.

Sensitive Detection of SARS-CoV-2 Using a SERS-Based Aptasensor.

We developed a new surface-enhanced Raman scattering (SERS)-based aptasensor platform capable of quantifying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lysates with a high sensitivity. In this study, a spike protein deoxyribonucleic acid (DNA) aptamer was used as a receptor, and a self-grown Au nanopopcorn surface was used as a SERS detection substrate for the sensible detection of SARS-CoV-2. A quantitative analysis of the SARS-CoV-2 lysate was performed by monitoring the change in the SERS peak intensity caused by the new binding between the aptamer DNA released from the Au nanopopcorn surface and the spike protein in the SARS-CoV-2 virion. This technique enables detecting SARS-CoV-2 with a limit of detection (LoD) of less than 10 PFU/mL within 15 min. The results of this study demonstrate the possibility of a clinical application that can dramatically improve the detection limit and accuracy of the currently commercialized SARS-CoV-2 immunodiagnostic kit.

Rapid and sensitive multiplex molecular diagnosis of respiratory pathogens using plasmonic isothermal RPA array chip.

The current clinically available multiplex molecular diagnostic technologies are difficult to apply to onsite diagnostic platforms due to their large and sophisticated instrumentation, long amplification times and limited number of simultaneous detections. We developed a plasmonic isothermal recombinase polymerase amplification (RPA) array chip for rapid and sensitive multiplex molecular detection. The 3D plasmonic substrate composed of Au nanoparticles (NPs) on dense Au nanopillars (NPOP) showed highly enhanced plasmon-enhanced fluorescence (PEF) of RPA products with long DNA amplicons (~200 bp). The plasmonic 4-plex RPA array chip successfully detected bacterial DNA within 30 min and viral RNA within 40 min; the sensitivity of the plasmonic RPA array chip was comparable to or 10-fold higher than that of the 4-pelx liquid-phase RPA and 4-plex liquid-phase PCR techniques. Additionally, no cross-reactivity was observed. The 4-plex plasmonic RPA array chip was preliminary evaluated using clinical respiratory viral-positive nasopharyngeal swab samples. This approach enables rapid, sensitive and high-multiplex molecular detection and can be used in the realization of a simplified and miniaturized platform for onsite multiplex molecular diagnostics.

A Wearable Surface-Enhanced Raman Scattering Sensor for Label-Free Molecular Detection.

A wearable surface-enhanced Raman scattering (SERS) sensor has been developed as a patch type to utilize as a molecular sweat sensor. Here, the SERS patch sensor is designed to comprise a sweat-absorbing layer, which is an interface to the human skin, an SERS active layer, and a dermal protecting layer that prevents damage and contaminations. A silk fibroin protein film (SFF) is a basement layer that absorbs aqueous solutions and filtrates molecules larger than the nanopores created in the ß-sheet matrix of the SFF. On the SFF layer, a plasmonic silver nanowire (AgNW) layer is formed to enhance the Raman signal of the molecules that penetrated through the SERS patch in a label-free method. A transparent dermal protecting layer (DP) allows laser penetration to the AgNW layer enabling Raman measurement through the SERS patch without its detachment from the surface. The molecular detection capability and time-dependent absorption properties of the SERS patch are investigated, and then, the feasibility of its use as a wearable drug detection sweat sensor is demonstrated using 2-fluoro-methamphetamine (2-FMA) on the human cadaver skin. It is believed that the developed SERS patch can be utilized as various flexible and wearable biosensors for healthcare monitoring.

A cyclodextrin-decorated plasmonic gold nanosatellite substrate for selective detection of bipyridylium pesticides.

A cyclodextrin-decorated gold nanosatellite (AuNSL) substrate was developed as a surface-enhanced Raman scattering sensor for the selective sensing of bipyridylium pesticides such as paraquat (PQ), diquat (DQ), and difenzoquat (DIF). The AuNSL structure was fabricated via vacuum deposition of gold nanoparticles (AuNPs) on a gold nanopillar substrate, and a large density of hot-spots was formed for Raman signal enhancement. Thiolated ß-cyclodextrin (SH-CD) was surface-modified on the AuNSL as a chemical receptor. The detection limit of PQ, DQ, and DIF on the SH-CD-coated AuNSL (CD-AuNSL) was 0.05 ppm for each, and showed linear correlation in a concentration range of 10 ppm-0.05 ppm. Then, selective bipyridylium pesticide detection was performed by comparing the Raman intensity of each pesticide with and without the washing step. After the washing step, 90% of the PQ, DQ, and DIF Raman signals were maintained on the CD-AuNSL substrate with a uniform selectivity in a mapping area of 200 µm × 200 µm. Furthermore, selective pesticide detection was performed using a ground-apple solution without pretreatment. Raman signals were clearly observed after the washing step and they showed a limit of detection down to a concentration of 0.05 ppm for each pesticide. Principal component analysis (PCA) of the binary and ternary mixtures of PQ, DQ, and DIF showed that each component could be easily identified via the typical Raman fingerprint analysis. The developed CD-AuNSL is expected to be applied for various chemical sensors, especially for pyridine-containing toxic substances in the environment and metabolite biomarkers in biofluids.

Plasmonic Microgels for Raman-Based Molecular Detection Created by Simultaneous Photoreduction and Photocross-linking.

Molecular detection in complex mixtures is of great importance in biomedical diagnosis, food safety, and environmental monitoring. Although surface-enhanced Raman scattering serves as one of the most promising detection methods, metal surfaces are prone to contamination, making the direct detection of small molecules in mixtures elusive. Metal nanoparticle-loaded hydrogels have been used for the exclusion of large adhesive molecules and direct detection of small molecules. Here, we design microgels containing highly concentrated gold nanoparticles through the simultaneous formation of hydrogel and gold nanoparticles in emulsion droplets. Monodisperse water-in-oil droplets are microfluidically prepared to contain a gold precursor, hydrogel precursor, and photoinitiator. Upon ultraviolet irradiation, a hydrogel is gradually formed in the drop by photocross-linking at which gold nanoparticles are synthesized and grown by photo and thermal reduction. The in situ synthesis provides the uniform distribution of gold nanoparticles at very high concentrations without aggregation, which is otherwise very difficult to achieve. Using the microgels, small molecules in albumin solutions can be detected by Raman measurement with high signal sensitivity and reproducibility in the absence of interruption from albumin. As a proof of concept, we demonstrate the direct detection of pyocyanin, a biomarker for Pseudomonas infection spiked in unpurified saliva.

SERS imaging-based aptasensor for ultrasensitive and reproducible detection of influenza virus A.

Surface-enhanced Raman scattering (SERS)-based aptasensors display high sensitivity for influenza A/H1N1 virus detection but improved signal reproducibility is required. Therefore, in this study, we fabricated a three-dimensional (3D) nano-popcorn plasmonic substrate using the surface energy difference between a perfluorodecanethiol (PFDT) spacer and the Au layer. This energy difference led to Au nanoparticle self-assembly; neighboring nanoparticles then created multiple hotspots on the substrate. The localized surface plasmon effects at the hot spots dramatically enhanced the incident field. Quantitative evaluation of A/H1N1 virus was achieved using the decrease of Raman peak intensity resulting from the release of Cy3-labeled aptamer DNAs from nano-popcorn substrate surfaces via the interaction between the aptamer DNA and A/H1N1 virus. The use of a Raman imaging technique involving the fast mapping of all pixel points enabled the reproducible quantification of A/H1N1 virus on nano-popcorn substrates. Average ensemble effects obtained by averaging all randomly distributed hot spots mapped on the substrate made it possible to reliably quantify target viruses. The SERS-based imaging aptasensor platform proposed in this work overcomes the issues inherent in conventional approaches (the time-consuming and labor-intensiveness of RT-PCR and low sensitivity and quantitative analysis limits of lateral flow assay kits). Our SERS-based assay for detecting A/H1N1 virus had an estimated limit of detection of 97 PFU mL-1 (approximately three orders of magnitude more sensitive than that determined by the enzyme-linked immunosorbent assay) and the approximate assay time was estimated to be 20 min. Thus, this approach provides an ultrasensitive, reliable platform for detecting viral pathogens.

Encapsulation of 3D plasmonic nanostructures with ultrathin hydrogel skin for rapid and direct detection of toxic small molecules in complex fluids.

Nanogap-rich 3D plasmonic nanostructures provide enhanced molecular Raman fingerprints in a nondestructive and label-free manner. However, the molecular detection of small target molecules in complex fluids is challenging due to nonspecific protein adsorption, which prevents access of the target molecules. Therefore, the molecular detection for complex mixtures usually requires a tedious and time-consuming pretreatment of samples. Herein, we report the encapsulation of 3D plasmonic nanostructures with an ultrathin hydrogel skin for the rapid and direct detection of small molecules in complex mixtures. To demonstrate the proof of concept, we directly detect pesticide dissolved in milk without pretreatment. This detection is enabled by the selective permeation of target molecules into the 3D mesh of the hydrogel skin and the adsorption onto plasmonic hotspots, accompanied by the rejection of large adhesive proteins and colloids. The high sensitivity of nanogap-rich plasmonic nanostructures in a conjunction with the molecular selection of the hydrogel skin enables the fast and reliable detection of tricyclazole in whole milk with a limit of detection as low as 10 ppb within 1 h. We believe that this plasmonic platform is highly adaptable for in situ and on-site detection of small molecules in various complex mixtures including foods, biological fluids, and environmental fluids.