Colloidal Gold Nanoparticles Research Articles

Lights on the Synthesis of Surfactant-Free Colloidal Gold Nanoparticles in Alkaline Mixtures of Alcohols and Water.

Surfactant-free colloidal syntheses in aqueous media are attractive to develop nanomaterials relevant for various applications, e. g. catalysis or medicine. However, controlled green syntheses without surfactants of metal nanoparticles in aqueous media remain scarce. Here, room temperature syntheses of gold (Au) nanoparticles (NPs) that require only HAuCl4, alkaline water and an alcohol, i. e. relatively benign chemicals and mild reaction conditions, are developed. The findings of a comprehensive multi-parameters screening performed in small volumes (<3 mL) over 1000+ experiments pave the way to greener high throughput screenings of large parametric spaces and lead to scalable (100 mL) synthetic strategies. A rational selection of the alcohol is proposed. The influence of lights with defined wavelengths (222-690 nm) is investigated. It is found that lights with lower wavelengths favor the formation of smaller 5 nm NPs. Different kinetics and formation pathways are observed for different alcohols and for lights with different wavelengths. The sensitivity to various experimental parameters increases with the alcohol used in the order glycerol<ethylene glycol<ethanol<methanol. New strategies for a rational fine size control, and to some extend shape control, are identified. The results lead to more sustainable and reproducible surfactant-free colloidal syntheses of NPs.

Spectrum reconstruction of experimentally produced nano-colloids using Mie theory

Aim: Synthesis of plasmonic nanoparticles, characterization, size detection by modeling. Methods: Colloidal plasmonic gold and silver nanoparticles were prepared by laser ablation with a 1,064 nm pulsed Nd:YAG laser with equal fractional volume and then were illuminated with its 532 nm second harmonic pulse. After the illumination process, we observed 1 nm blue shift in peak position of gold colloid and 4 nm blue shift in silver colloid. We observed a variation in the dielectric function due to nanoparticle size reduction in both samples. Using micrograph, size distribution was plotted and with the help of Mie theory and size dependent dielectric function, we reconstructed absorption spectrum to best fit the experimental spectrum and we estimated 12- and 16-fold increase in the number of Au and Ag nanoparticles respectively, due to illumination. Results: We have estimated size distribution of produced nanoparticles. Conclusions: We produced silver and gold colloids with ablation of their foils in water without any surfactant, and then we fragmented the nanoparticles colloids with an intense nanosecond laser and studied the effect of illumination on peak position and size distribution of colloids.

Defect-Selective Energy Barrier Crossing during Adsorption of Colloidal Gold Nanoparticles on Zinc Sulfide Crystals under Overall Electrostatic Repulsion

Defect-Selective Energy Barrier Crossing during Adsorption of Colloidal Gold Nanoparticles on Zinc Sulfide Crystals under Overall Electrostatic Repulsion

Facile Electrical DNA Genosensor for Human Papillomavirus (HPV 58) for Early Detection of Cervical Cancer

For decades, a Pap smear test has been applied as a conventional method in detecting Human Papillomavirus caused cervical cancer. False-positive results were also recorded while using it as conventional method. Current biosensor such as Hybrid (II) Capture resulted in higher time consumption and cost. s Meanwhile, in this study we provided facile, mini, rapid, highly sensitive, eco-friendly, and cost-effective sensing system focusing on HPV strain 58 (HPV58) in a nano-size lab-on-chip technology genosensor. 30-mer of virus ssDNA designed and analyzed as a probe via bioinformatics tools such as GenBank, Basic Local Alignment Searching Tools (BLAST) and ClustalW. Nanotechnology-developed colloidal Gold-nanoparticles (AuNPs) are used in the biosensor fabrication to produce high stability and electron efficient transmission during electrical measurement. AuNPs-APTES modified on active sites of IDEs, followed by immobilization of specific probe ssDNA for HPV 58. Hydrogen binding during hybridization with its target produce electrical signals measured by KEITHLEY 2450 (Source Meter). The genosensor validated with different types of targets such as complimentary, non-complementary and single mismatch oligonucleotides. The serial dilution of target concentration has been experimented triplicate (n=3) range from 1fM to 10μM. The slope of calibration curve resulted 2.389E-0 AM-1 with regression coefficient (R2) = 0.97535.

UV light promoted dihydrolipoic acid and its alanine derivative directed rapid synthesis of stable gold nanoparticles and their catalytic activity

In general, colloidal gold nanoparticles (AuNPs) have been synthesized in heated or boiling water containing HAuCl4 precursor with sodium citrate as reducing stabilizing reagent. Although temperature plays a driving for synthesis of AuNPs, elevated temperature in thermal reduction method causes aggregation of the AuNPs. The preferential, rapid and strong binding of dihydro-lipoic acid and its derivatives on surface of AuNPs via thiol − Au chemistry promote the production of very stable AuNPs. In this study, we have developed citric acid (CA), dihydrolipoic acid (DHLA) and DHLA-Alanine (DHLA-Ala) directed rapid synthesis of ultra-stable AuNPs, DHLA@AuNPs and DHLA-Ala@AuNPs, under the UV (311 nm) irradiation at room temperature (RT: 25 °C) in around 10 min (min). CA is used as a potential reducing agent to expedite both reduction of Au3+ ion and AuNP formation, DHLA and DHLA-Ala act as stabilizing agents by replacing CA molecules on surface of AuNPs in order to produce quite stable AuNP. It is worthy to mention that reduction of Au3+ ion, formation and surface stabilization of AuNPs are consequently occurred in one step. We also investigated how experimental parameters including reaction time and temperature, pH of reaction solution, affect formation of the AuNPs. The effects of salt concentration and storage temperature were studied to show stability of the AuNPs. The synthesized DHLA@AuNPs and DHLA-Alanine@AuNPs were characterized via UV-Vis spectrophotometer (UV-Vis), scanning transmission electron microscope (STEM), dynamic light scattering (DLS) and Zeta potential (ZT) devices. The reduction of 4-nitrophenol (4-NP) to 4-aminophenol (4-AP) was efficiently catalyzed by the AuNPs in the presence of sodium borohydride in aqueous solution.

Surface Functionalization of Gold Nanoparticles Using Alkyne Derivatives: Applications in Chemical Sensing.

Colloidal gold nanoparticles (AuNPs) are important nanomaterials for chemical sensing and therapeutics. For their application, it is vital to develop a reliable and robust surface functionalization method that can be applied to diverse functional molecules and offer better stability under harsh biological conditions. Currently, thiol (SH) is the most commonly used functional group for forming stable covalent bonds with AuNPs. However, thiolated molecules typically require complicated preparation procedures, are susceptible to oxidation, and are not compatible with many electrophiles and reducing groups. In this study, we report that surface functionalization of AuNPs can be achieved using alkyne derivatives, which exhibit several advantages over classical thiolation and peptide-bond methods, including straightforward preparation of alkyne derivatives, rapid and simple conjugation in buffers and complex media, higher conjugation efficiency, long-term stability, and resistance to decomposition under harsh conditions. Several alkynylated biotin and fluorescein derivatives are prepared, and the alkynylated-AuNPs are characterized using a lateral flow assay, gel electrophoresis, and spectroscopy techniques to investigate the conjugation efficiencies, size distributions, protein interaction properties, and binding mode of the Au-alkyne bond. We also demonstrate that alkynylated-AuNPs can be used for the sensitive detection of hydrogen peroxide and streptavidin proteins.

Dynamic X-ray Coherent Diffraction Analysis: Bridging the Time Scales between Imaging and Photon Correlation Spectroscopy.

The advent of diffraction limited sources and developments in detector technology opens up new possibilities for the study of materials in situ and operando. Coherent X-ray diffraction techniques such as coherent X-ray diffractive imaging (CXDI) and X-ray photon correlation spectroscopy (XPCS) are capable for this purpose and provide complementary information, although due to signal-to-noise requirements, their simultaneous demonstration has been limited. Here, we demonstrate a strategy for the simultaneous use of CXDI and XPCS to study in situ the Brownian motion of colloidal gold nanoparticles of 200 nm diameter suspended in a glycerol-water mixture. We visualize the process of agglomeration, examine the spatiotemporal space accessible with the combination of techniques, and demonstrate CXDI with 22 ms temporal resolution.

Low-Temperature Sintering of Colloidal Gold Nanoparticles by Salt Addition

AbstractGold nanoparticles synthetized by pulsed laser ablation in liquid with a mean diameter of 4 nm were joined together by adding potassium bromide solution at various concentrations. By increasing the salt concentration, there is a significant increase of the particle size up to a mean diameter of 18 nm. We have studied the nanoparticle merging by using atomic force and electron microscopy characterizations, also demonstrating that it is possible to deposit sintered nanoparticles on silanized substrates in a fast, simple, cost-effective, energy-saving method with relevance in industrial manufacturing.

Synthesis and Characterization of Acacia-Stabilized Doxorubicin-Loaded Gold Nanoparticles for Breast Cancer Therapy.

The targeted delivery of drugs is vital in breast cancer treatment due to its ability to produce long-lasting therapeutic effects with minimal side effects. This study reports the successful development of doxorubicin hydrochloride (DOX)-loaded colloidal gold nanoparticles stabilized with acacia gum (AG). Optimization studies varied AG concentrations (0.25% to 3% w/v) to determine optimal conditions for nanoparticle synthesis. The resulting acacia stabilized gold nanoparticles (AGNPs) were characterized using various techniques including high-resolution transmission electron microscopy (HR-TEM), powder X-ray diffraction (PXRD), differential scanning calorimetry (DSC), ultraviolet-visible spectroscopy, Fourier-transform infrared spectroscopy (FT-IR), field emission scanning electron microscopy (FE-SEM), and selected area electron diffraction (SAED). In vitro drug release studies demonstrated a higher release rate of DOX in sodium acetate buffer (pH 5.0) compared to phosphate buffer saline (pH 7.4), suggesting an enhanced therapeutic efficacy in acidic tumor environments. Cytotoxicity of DOX-AGNPs and free DOX was assessed in human breast cancer cells (MDA-MB-231). The DOX-AGNPs exhibited significantly greater cytotoxicity, indicating enhanced efficacy in targeting cancer cells. This enhancement suggests that adsorbing DOX on the surface of gold nanoparticles can improve drug delivery and effectiveness, potentially reducing side effects compared to pure DOX and traditional delivery methods. Stability tests conducted over six months at 25±1°C showed significant changes in particle size and PDI, suggesting limited stability under these conditions. Overall, the acacia-stabilized gold nanoparticles synthesized in this study exhibit promising characteristics for drug delivery applications, particularly in cancer therapy, with effective drug loading, controlled release, and favorable physicochemical properties.

Ultrasensitive Rapid Antigen Test by Geometric Lateral Flow Assays and Highly Doped Upconversion Nanoparticles.

The paper-based lateral flow assay (LFA) testing strips are currently the most widely used for point-of-care testing (POCT), valued for their rapid result turnaround times in a few minutes. However, their sensitivity has been limited. Upconversion nanoparticles (UCNPs), especially highly doped ones, have emerged as promising luminescent reporters to enhance the LFA sensitivity. These UCNPs exhibit a nonlinear enhancement in luminescence with excitation power density, necessitating higher power densities for higher brightness. In this study, we utilized a geometric paper strip design to minimize the immune reaction area and maximize the excitation power density, enabling ultrasensitive detection of the SARS-CoV-2 nucleoprotein antigen. This design also slowed the antigen flow on the paper strip, extending the reaction time between antigen and antibody, thereby enhancing the efficiency of the immune reaction. Through this design, our approach achieved over a 100-fold enhancement in the limit of detection (LOD) compared with the widely used LFAs, based on gold colloidal nanoparticles and europium nanoparticles. This innovation expands the scope of LFA applications that require a low LOD.

Surfactant-Free Method to Prevent Gold Nanoparticle Aggregation and Its Surface-Enhanced Raman Scattering Applications.

The introduction of surfactants to stabilize colloidal citrate-reduced gold nanoparticles (prevent aggregation) is usually used in surface-enhanced Raman scattering (SERS) applications. However, the surfactants have many drawbacks for SERS applications, such as increasing the SERS background and blocking surface active sites. Here, we develop a surfactant-free method to stabilize colloidal cit-AuNPs based on alkali regulation, and this method can prevent gold nanoparticle aggregation under different harsh treatments, including ligand modification, centrifugation-based washing/enrichment, and salt addition. The SERS spectra, density functional theory simulation, and ζ potentials of cit-AuNPs indicate that the stability of enhanced cit-AuNPs under alkaline conditions is attributed to both the increased negative charge density (by ∼6 times from pH 7 to 12) and the molecular configuration on the metal surface. Compared with surfactant-based methods, this method can well maintain the inherent optical and interface properties of nanoparticles, avoid the SERS background, and avoid blocking of the surface active site due to the presence of surfactants. This method will enable AuNPs to have a wide range of applications in areas such as highly sensitive SERS sensors.

Application of stealth nanobeacon for traceability assurance in pharmaceutical tablets via surface-enhanced Raman scattering during the tablet coating process

Microtaggant technologies for on-dose authentication have garnered significant interest for use in the anti-counterfeit activities and traceability of pharmaceutical dosage forms. Previously, we proposed a stealth nanobeacon (NB) comprising self-assembled colloidal gold nanoparticles with reporter molecules that demonstrated characteristic surface-enhanced Raman scattering (SERS) activity. However, the integration of such microtaggants into standard production lines remains underexplored. In this study, we demonstrate the incorporation of NB into tablet coatings using a simple mixing method with conventional coating solutions. Rapid and discernible SERS responses from the NB-coated tablets were observed in response to laser excitation at 785 nm for 0.1s, implying that it is an advanced and efficient method for counterfeit detection. In addition, the SERS intensity of NB increased with coating time, suggesting that NB can be used as a tracer for the real-time monitoring of coating thickness. Furthermore, NB-coated tablets were indistinguishable from NB-free tablets, even during colorimetric analysis. These results suggest that the NB possesses stealth properties and can be easily incorporated into counterfeit detection products.

Chiral sensing of glucose by surface-enhanced Raman spectroscopy

BackgroundChiral-selective molecular interactions are considered crucial in numerous physiological processes. Chiral-selective analytical methods of biomolecules with sufficient sensitivity are of great interest in numerous applications. Several surface-enhanced Raman scattering (SERS)-based methods have recently been reported for chiral sensing of biomolecules. However, the lack of molecular-level understanding of SERS spectral changes of reporter and analyte molecules may mislead the development of chiral detection methods. ResultsWe report the chiral sensing of glucose (Glu) by SERS of L- and D-phenylalanine (Phe) with colloidal gold nanoparticles (AuNPs) synthesized by borohydride ions. The Phe SERS showed drastic spectral changes only when Glu of the same chirality as Phe was added, which also showed strong dependence on Glu concentration. The increase of δ(COO−) and decrease of νs(COO−) modes in Phe SERS, exclusively observed with the chiral-selective bimolecular interactions of chirally matching Glu, are understood as modified surface adsorption geometry of the carboxylate group. Quantitative spectral analysis for the Glu concentration of a specific chirality showed the detection limit down to 2 × 10−9 - 2 × 10−7 M levels depending on the existence of the opposite enantiomer of Glu. SignificanceIn this study, we demonstrated that the Phe SERS on AuNPs can be utilized in the chiral sensing of Glu molecules with quantitative concentration analysis. The bimolecular interactions of surface-adsorbed Phe and chirally matching Glu are suggested for the chiral recognition of Phe SERS. These results imply that a molecule-level understanding is indispensable for developing SERS-based chiral sensing methods.

Differential detection of SARS-CoV-2 variants and influenza A viruses utilizing a dual lateral flow strip based on colloidal gold-labeled monoclonal antibodies

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the seasonal influenza virus are spreading among humans concurrently, especially with the ongoing replacement of mutant strains. It is challenging to differentiate between symptoms for therapy due to the comparable symptoms following infection with the SARS-CoV-2 variants and influenza viruses. Meanwhile, in order to achieve rapid point-of-care testing (POCT) to manage the spread of the disease, we developed a dual lateral flow strip based on colloidal gold-labeled monoclonal antibodies that can perform differential detection of SARS-CoV-2 variants and influenza A viruses (IAV) in this study. High-affinity monoclonal antibodies (mAbs) targeting SARS-CoV-2 and IAV were prepared to capture antigens and labeled with colloidal gold nanoparticles (AuNPs). Based on high-affinity mAbs, two targets were immobilized on one nitrocellulose (NC) membrane to establish the lateral flow strip (LFS) for differential diagnosis of SARS-CoV-2 and IAV. With no reactivity to other viruses, this LFS is extremely specific and can only identify SARS-CoV-2 and IAV. The LFS showed a limit of detection (LOD) of 4.88 ng/mL for the Omicron BA.2 RBD protein and 2.44 ng/mL for the nucleoprotein (NP) protein of H1N1. When analyzing 16 SARS-CoV-2 positive clinical samples, eight IAV positive clinical samples, and six negative samples that had already been pre-confirmed by commercial kits, its clinical application is effectively and accurately proven. These results demonstrated that the LFS integrated with AuNPs has great potential to facilitate quick, easy, and reliable POCT diagnosis for promoting the control of infectious diseases.

Integrated Ultrasound-Enrichment and Machine Learning in Colorimetric Lateral Flow Assay for Accurate and Sensitive Clinical Alzheimer's Biomarker Diagnosis.

The colloidal gold nanoparticle (AuNP)-based colorimetric lateral flow assay (LFA) is one of the most promising analytical tools for point-of-care disease diagnosis. However, the low sensitivity and insufficient accuracy still limit its clinical application. In this work, a machine learning (ML)-optimized colorimetric LFA with ultrasound enrichment is developed to achieve the sensitive and accurate detection of tau proteins for early screening of Alzheimer's disease (AD). The LFA device is integrated with a portable ultrasonic actuator to rapidly enrich microparticles using ultrasound, which is essential for sample pre-enrichment to improve the sensitivity, followed by ML algorithms to classify and predict the enhanced colorimetric signals. The results of the undiluted serum sample testing show that the protocol enables efficient classification and accurate quantification of the AD biomarker tau protein concentration with an average classification accuracy of 98.11% and an average prediction accuracy of 99.99%, achieving a limit of detection (LOD) as sensitive as 10.30pgmL-1. Further point-of-care testing (POCT) of human plasma samples demonstrates the potential use of LFA in clinical trials. Such a reliable lateral flow immunosensor with high precision and superb sensing performance is expected to put LFA in perspective as an AD clinical diagnostic platform.

AFM for Nanomechanical Assessment of Polymer Overcoatings on Nanoparticle-Decorated Biomaterials.

Nanoparticle adhesion to polymer and similar substrates may be prone to low nano-Newton forces, disrupting the surface bonds and patterning, potentially reducing the functionality of complex surface patterns. Testing this, a functionalised surface reported for biological and medical applications, consisting of a thin plasma-derived oxazoline-based film with 68 nm diameter covalently bound colloidal gold nanoparticles attached within an aqueous solution, underwent nanomechanical analysis. Atomic Force Microscopy nanomechanical analysis was used to quantify the limits of various adaptations to these nanoparticle-featured substrates. Regular and laterally applied forces in the nano-Newton range were shown to de-adhere surface-bound gold nanoparticles. Applying a nanometre-thick overcoating anchored the nanoparticles to the surface and protected the underlying base substrate in a one-step process to improve the overall stability of the functionalised substrate against lower-range forces. The thickness of the oxazoline-based overcoating displayed protection from forces at different rates. Testing overcoating thickness ranging from 5 to 20 nm in 5 nm increments revealed a significant improvement in stability using a 20 nm-thick overcoating. This approach underscores the importance of optimising overcoating thickness to enhance nanoparticle-based surface modifications' durability and functional integrity.

Quantitative and Ultrasensitive Detection of Four Inflammatory Biomarkers by Gold Nanoparticle‐Based Paper Sensor

AbstractDue to the complexity and diversity of infectious diseases, diagnosis based solely on symptoms and imaging manifestations is associated with a high rate of misdiagnosis, so it is particularly important to find reliable and accurate biomarkers. Four inflammatory biomarkers (CRP, IL‐6, PCT and SAA) have different manifestations and roles in different infectious diseases and stages of infection, and the diagnosis and identification of infectious diseases can be realized through quantitative analysis of these four biomarkers simultaneously. In this study, we developed a colloidal gold nanoparticle (AuNP)‐based multiplex LFA strip using eight highly‐sensitive monoclonal antibodies that simultaneously detected CRP, IL‐6, PCT and SAA in serum within 15 min. And quantitative analysis was achieved with a portable strip reader, the LODs were 2.5 μg/mL, 25.3 pg/mL 0.87 ng/mL and 8.8 μg/mL for CRP, IL‐6, PCT and SAA, while detection ranges were 2.5–200 μg/mL, 25.3–8000 pg/mL 0.87–100 ng/mL and 8.8–200 μg/mL. Moreover, the quantitative colloidal gold assay correlated well with the results of a chemiluminescence immunoassay when testing clinical serum samples. Our developed method was reliable and accurate according to the recovery test results. Therefore, the strip can be used as an alternative method for the simultaneous monitoring of CRP, IL‐6, PCT and SAA in serum samples.

Hierarchical magneto-colorimetric labels for immediate lateral flow immunoassay of chlorothalonil residues

Quantitative detection of pesticide residues in food and environmental samples using an improved lateral flow immunoassay (LFIA) is of considerable importance for real-time analysis. This paper proposes a highly sensitive LFIA platform based on a hierarchical magneto-colorimetric compact. This compact serves as both the target magnetic enrichment substrate and a photosensitive label. Initially, a large porous dendritic silica template is prepared and doped with superparamagnetic ferric oxide nanoparticles (Fe3O4 NPs) and colloidal gold nanoparticles (AuNPs) at high densities within its vertical channels. The sequential assembly of central–radial channels allow for the three-dimensional integration of these two components, enabling independent control of their discrete functions without mutual interference. Following alkyl organosilicon encapsulation and silica sealing, the composite spheres are then applied in LFIA to detect chlorothalonil residues. Fe3O4 NPs enhance the binding efficiency to target analytes, while AuNPs amplify the signal, leveraging their high loading densities and robust optical properties. The developed LFIA platform exhibited a detection limit of 0.34 ng/mL for chlorothalonil and a linear range of 0.0085–824 ng/mL. The recoveries varied between 85.1 % and 103.1 %, and the relative standard deviations were 1.25%–8.84 %. This LFIA approach demonstrates high sensitivity, specificity, reproducibility and flexible detection modes, making it highly suitable for the on-site monitoring of pesticide residues.

Self-Regulated Assembly and Disassembly of Gold Nanoparticles for Low-Temperature Time Indication.

The color-changing self-assembly and autonomous disassembly of colloidal gold nanoparticles (AuNPs) is reported by simply mixing negatively charged phosphine ligand-capped AuNPs with partially oxidized polyethylene glycol (PEG). The assembly of AuNPs is initiated by PEG adsorption, which disrupts the hydration layer of AuNPs, leading to depletion attraction and reduction of hydration repulsion among the AuNPs. The oxidative species in PEG subsequently oxidize and remove the charged ligands from the AuNP surface, resulting in a decrease and reversal of the negative surface charge. This causes the PEG to adsorb on AuNPs in a tighter and more direct manner, providing strong steric shielding to the AuNPs, thereby triggering the disassembly of the AuNP assemblies. The self-regulated assembly-disassembly process can be tuned widely by controlling chemical conditions of PEG, nanoparticle concentration, and the environmental conditions, suggesting potential applications as colorimetric time-temperature indicators for food and medicine storage conditions. As a proof of concept, it is demonstrated that the lifetime of the color-changing assembly-disassembly process can be extended from tens of minutes to weeks when subjected to a refrigerated environment, with tunability achievable through varying polymer conditions and storage atmospheres.

Direct Writing of SERS Substrates Using Femtosecond Laser Pulses.

Achieving a high-density, repeatable, and uniform distribution of "hotspots" across the entire surface-enhanced Raman scattering (SERS) substrate is a current challenge in facilitating the efficient preparation of large-area SERS substrates. In this study, we aim to produce homogeneous surface-enhanced Raman scattering (SERS) substrates based on the strong interaction between femtosecond laser pulses and a thin film of colloidal gold nanoparticles (AuNPs). The SERS substrate we obtained consists of irregularly shaped and sharp-edged gold nanoparticle aggregates with specially extruding features; meanwhile, a large number of three-dimensional AuNP stacks are produced. The advantages of such configurations lie in the production of a high density of hotspots, which can significantly improve the SERS performance. When the laser fluence is 5.6 mJ/cm2, the substrate exhibits the best SERS enhancement effect, and a strong SERS signal can still be observed when testing the concentration of R6G at 10-8 mol/L. The enhancement factor of such SERS substrates prepared using femtosecond laser direct writing is increased by 3 orders of magnitude compared to the conventional furnace annealing process. Furthermore, the relative standard deviation for the intensities of the SERS signals was measured to be 5.1% over an area of 50 × 50 μm2, indicating a highly homogeneous SERS performance and excellent potential for practical applications.