Effects of Raman Labeling Compounds on the Stability and Surface-Enhanced Raman Spectroscopy Performance of Ag Nanoparticle-Embedded Silica Nanoparticles as Tagging Materials.

Surface-enhanced Raman spectroscopy (SERS) tagging using silica(SiO2)@Ag nanoparticles (NPs) is easy to handle and is being studied in various fields, including SERS imaging and immunoassays. This is primarily due to its structural advantages, characterized by high SERS activity. However, the Ag NPs introduced onto the SiO2 surface may undergo structural transformation owing to the Ostwald ripening phenomenon under various conditions. As a result, the consistency of the SERS signal decreases, reducing their usability as SERS substrates. Until recently, research has been actively conducted to improve the stability of single Ag NPs. However, research on SiO2@Ag NPs used as a SERS-tagging material is still lacking. In this study, we utilized a Raman labeling compound (RLC) to prevent the structural deformation of SiO2@Ag NPs under various conditions and proposed excellent SiO2@Ag@RLC-Pre NPs as a SERS-tagging material. Using various RLCs, we confirmed that 4-mercaptobenzoic acid (4-MBA) is the RLC that maintains the highest stability for 2 months. These results were also observed for the SiO2@Ag NPs, which were unstable under various pH and temperature conditions. We believe that SERS tags using SiO2@Ag NPs and 4-MBA can be utilized in various applications on based SERS because of the high stability and consistency of the resulting SERS signal.

Optical nanomaterial-based detection of biomarkers in liquid biopsy.

Liquid biopsy, which is a minimally invasive procedure as an alternative to tissue biopsy, has been introduced as a new diagnostic/prognostic measure. By screening disease-related markers from the blood or other biofluids, it promises early diagnosis, timely prognostication, and effective treatment of the diseases. However, there will be a long way until its realization due to its conceptual and practical challenges. The biomarkers detected by liquid biopsy, such as circulating tumor cell (CTC) and circulating tumor DNA (ctDNA), are extraordinarily rare and often obscured by an abundance of normal cellular components, necessitating ultra-sensitive and accurate detection methods for the advancement of liquid biopsy techniques. Optical biosensors based on nanomaterials open an important opportunity in liquid biopsy because of their enhanced sensing performance with simple and practical properties. In this review article, we summarized recent innovations in optical nanomaterials to demonstrate the sensitive detection of protein, peptide, ctDNA, miRNA, exosome, and CTCs. Each study prepares the optical nanomaterials with a tailored design to enhance the sensing performance and to meet the requirements of each biomarker. The unique optical characteristics of metallic nanoparticles (NPs), quantum dots, upconversion NPs, silica NPs, polymeric NPs, and carbon nanomaterials are exploited for sensitive detection mechanisms. These recent advances in liquid biopsy using optical nanomaterials give us an opportunity to overcome challenging issues and provide a resource for understanding the unknown characteristics of the biomarkers as well as the mechanism of the disease.

Recent Studies on Metal-Embedded Silica Nanoparticles for Biological Applications.

Recently, silica nanoparticles (NPs) have attracted considerable attention as biocompatible and stable templates for embedding noble metals. Noble-metal-embedded silica NPs utilize the exceptional optical properties of novel metals while overcoming the limitations of individual novel metal NPs. In addition, the structure of metal-embedded silica NPs decorated with small metal NPs around the silica core results in strong signal enhancement in localized surface plasmon resonance and surface-enhanced Raman scattering. This review summarizes recent studies on metal-embedded silica NPs, focusing on their unique designs and applications. The characteristics of the metal-embedded silica NPs depend on the type and structure of the embedded metals. Based on this progress, metal-embedded silica NPs are currently utilized in various spectroscopic applications, serving as nanozymes, detection and imaging probes, drug carriers, photothermal inducers, and bioactivation molecule screening identifiers. Owing to their versatile roles, metal-embedded silica NPs are expected to be applied in various fields, such as biology and medicine, in the future.

High-Throughput Synthesis of Nanogap-Rich Gold Nanoshells Using Dual-Channel Infusion System.

Gold nanoshells have been actively applied in industries beyond the research stage because of their unique optical properties. Although numerous methods have been reported for gold nanoshell synthesis, the labor-intensive and time-consuming production process is an issue that must be overcome to meet industrial demands. To resolve this, we report a high-throughput synthesis method for nanogap-rich gold nanoshells based on a core silica support (denoted as SiO2@Au NS), affording a 50-fold increase in scale by combining it with a dual-channel infusion pump system. By continuously dropping the reactant solution through the pump, nanoshells with closely packed Au nanoparticles were prepared without interparticle aggregation. The thickness of the gold nanoshells was precisely controlled at 2.3-17.2 nm by regulating the volume of the reactant solution added dropwise. Depending on the shell thickness, the plasmonic characteristics of SiO2@Au NS prepared by the proposed method could be tuned. Moreover, SiO2@Au NS exhibited surface-enhanced Raman scattering activity comparable to that of gold nanoshells prepared by a previously reported low-throughput method at the same reactant ratio. The results indicate that the proposed high-throughput synthesis method involving the use of a dual-channel infusion system will contribute to improving the productivity of SiO2@Au NS with tunable plasmonic characteristics.

Advanced Optical Materials: From Materials to Applications.

Optical materials interact significantly with electromagnetic radiation in the visible, ultraviolet, and infrared regions of the spectrum [...].

Mesoporous Silica Particle as an RNA Adsorbent for Facile Purification of In Vitro-Transcribed RNA.

Messenger RNA vaccines against SARS-CoV-2 hold great promise for the treatment of a wide range of diseases by using mRNA as a tool for generating vaccination antigens as well as therapeutic proteins in vivo. Increasing interest in mRNA preparation warrants reliable methods for in vitro transcription (IVT) of mRNA, which must entail the elimination of surplus side products such as immunogenic double-stranded RNA (dsRNA). We developed a facile method for the removal of dsRNA from in vitro transcribed RNA with mesoporous silica particles as RNA adsorbents. Various polyamines were tested for the facilitation of RNA adsorption onto mesoporous silica particles in the chromatography. Among the polyamines tested for RNA adsorption, spermidine showed a superior capability of RNA binding to the silica matrix. Mesoporous silica-adsorbed RNA was readily desorbed with elution buffer containing either salt, EDTA, or urea, possibly by disrupting electrostatic interaction and hydrogen bonding between RNA and the silica matrix. Purification of IVT RNA was enabled with the adsorption of RNA to mesoporous silica in a spermidine-containing buffer and subsequent elution with EDTA. By differing EDTA concentration in the eluting buffer, we demonstrated that at least 80% of the dsRNA can be removed from the mesoporous silica-adsorbed RNA. When compared with the cellulose-based removal of dsRNA from IVT RNA, the mesoporous silica-based purification of IVT RNA using spermidine and EDTA in binding and elution, respectively, exhibited more effective removal of dsRNA contaminants from IVT RNA. Thus, mRNA purification with mesoporous silica particles as RNA adsorbents is applicable for the facile preparation of nonimmunogenic RNA suitable for in vivo uses.

Recent Trends in Lateral Flow Immunoassays with Optical Nanoparticles.

Rapid, accurate, and convenient diagnosis is essential for effective disease management. Various detection methods, such as enzyme-linked immunosorbent assay, have been extensively used, with lateral flow immunoassay (LFIA) recently emerging as a major diagnostic tool. Nanoparticles (NPs) with characteristic optical properties are used as probes for LFIA, and researchers have presented various types of optical NPs with modified optical properties. Herein, we review the literature on LFIA with optical NPs for the detection of specific targets in the context of diagnostics.

Functional Optical Nano/Micromaterials.

The interaction between light and optical materials is central to science, as these materials possess remarkable physical, chemical, and photonical characteristics [...].

In Vitro Tracking of Human Umbilical Vein Endothelial Cells Using Ultra-Sensitive Quantum Dot-Embedded Silica Nanoparticles.

The nanoscale spatiotemporal resolution of single-particle tracking (SPT) renders it a powerful method for exploring single-molecule dynamics in living cells or tissues, despite the disadvantages of using traditional organic fluorescence probes, such as the weak fluorescent signal against the strong cellular autofluorescence background coupled with a fast-photobleaching rate. Quantum dots (QDs), which enable tracking targets in multiple colors, have been proposed as an alternative to traditional organic fluorescence dyes; however, they are not ideally suitable for applying SPT due to their hydrophobicity, cytotoxicity, and blinking problems. This study reports an improved SPT method using silica-coated QD-embedded silica nanoparticles (QD2), which represent brighter fluorescence and are less toxic than single QDs. After treatment of QD2 in 10 µg/mL, the label was retained for 96 h with 83.76% of labeling efficiency, without impaired cell function such as angiogenesis. The improved stability of QD2 facilitates the visualization of in situ endothelial vessel formation without real-time staining. Cells retain QD2 fluorescence signal for 15 days at 4 °C without significant photobleaching, indicating that QD2 has overcome the limitations of SPT enabling long-term intracellular tracking. These results proved that QD2 could be used for SPT as a substitute for traditional organic fluorophores or single quantum dots, with its photostability, biocompatibility, and superior brightness.

Synthesis and Applications of Optical Materials.

As optical materials have shown outstanding physical and chemical characteristics in the bio, medical, electronics, energy and related fields of studies, the potential benefits of using these materials have been widely recognized [...].

Efficient Analysis of Small Molecules via Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (LDI-TOF MS) Using Gold Nanoshells with Nanogaps.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a commonly used technique for analyzing large biomolecules. However, the utilization of organic matrices limits the small-molecule analysis because of the interferences in the low-mass region and the reproducibility issues. To overcome these limitations, a surface-assisted laser desorption/ionization (SALDI), which utilizes nanostructured metallic surfaces, has been developed. Herein, a novel approach for SALDI-MS was proposed using silica@gold core-shell hybrid materials with a nanogap-rich shell (SiO2@Au NGS), which is an emerging material due to its excellent heat-generating capabilities. The gold shell thickness was controlled by adjusting the concentration of gold precursor for the growth of gold nanoparticles. SALDI-MS measurements were performed on a layer formed by drop-casting a mixture of SiO2@Au NGS and analytes. At the optimized process, the gold shell thickness was observed to be 17.2 nm, which showed the highest absorbance. Based on the enhanced SALDI capability, SiO2@Au NGS was utilized to detect various small molecules, including amino acids, sugars, and flavonoids, and the ionization softness was confirmed with a survival yield upon fragmentation. The limits of detection, reproducibility, and salt tolerance of SiO2@Au NGS demonstrate its potential as an effective and reliable SALDI material for small-molecule analyses.

Mechanical Timer-Actuated Fluidic Dispensing System: Applications to an Automated Multistep Lateral Flow Immunoassay with High Sensitivity.

In this study, we present a fluidic dispensing system that can automate the sequential fluidic delivery of multiple reagents for lateral flow assays. Highly sensitive assays typically require multiple solution-based sequences, including washing steps and signal amplification. However, implementation of these types of sequences on an automated and highly sensitive point-of-care testing (POCT) platform remains challenging. Our platform consists of two disposable cartridges with reagent chambers and a test strip and an instrument that has a mechanical timer to actuate the cam-follower-gear components. The timer rotation sequentially shifts the position of the chambers and loads the reagents to the test paper strip. The dispensing intervals are controlled at a variation of <1% within a total actuation time of 60 min. Unlike other POCT devices, the timing of fluid delivery in our timer-actuated platform is not dependent on the selection of substrates and reagents, and the unique approach to fluidic delivery results in no reagent overlap or carryover, minimal reagent loss, and highly accurate fluidic timing control for highly sensitive solution-based assays. As a model application, the proposed platform applies a gold enhancement solution to amplify the detection signal and detect prostate-specific antigen with a limit of detection of 86 pg/mL within 27 min. This platform provides an opportunity for solution-based POCT applications with high sensitivity, thereby satisfying the requirement for user-friendly operations in resource-limited settings.

Silica Shell Thickness-Dependent Fluorescence Properties of SiO2@Ag@SiO2@QDs Nanocomposites.

Silica shell coatings, which constitute important technology for nanoparticle (NP) developments, are utilized in many applications. The silica shell's thickness greatly affects distance-dependent optical properties, such as metal-enhanced fluorescence (MEF) and fluorescence quenching in plasmonic nanocomposites. However, the precise control of silica-shell thicknesses has been mainly conducted on single metal NPs, and rarely on complex nanocomposites. In this study, silica shell-coated Ag nanoparticle-assembled silica nanoparticles (SiO2@Ag@SiO2), with finely controlled silica shell thicknesses (4 nm to 38 nm), were prepared, and quantum dots (QDs) were introduced onto SiO2@Ag@SiO2. The dominant effect between plasmonic quenching and MEF was defined depending on the thickness of the silica shell between Ag and QDs. When the distance between Ag NPs to QDs was less than ~10 nm, SiO2@Ag@SiO2@QDs showed weaker fluorescence intensities than SiO2@QD (without metal) due to the quenching effect. On the other hand, when the distance between Ag NPs to QDs was from 10 nm to 14 nm, the fluorescence intensity of SiO2@Ag@SiO2@QD was stronger than SiO2@QDs due to MEF. The results provide background knowledge for controlling the thickness of silica shells in metal-containing nanocomposites and facilitate the development of potential applications utilizing the optimal plasmonic phenomenon.

Highly Bright Silica-Coated InP/ZnS Quantum Dot-Embedded Silica Nanoparticles as Biocompatible Nanoprobes.

Quantum dots (QDs) have outstanding optical properties such as strong fluorescence, excellent photostability, broad absorption spectra, and narrow emission bands, which make them useful for bioimaging. However, cadmium (Cd)-based QDs, which have been widely studied, have potential toxicity problems. Cd-free QDs have also been studied, but their weak photoluminescence (PL) intensity makes their practical use in bioimaging challenging. In this study, Cd-free QD nanoprobes for bioimaging were fabricated by densely embedding multiple indium phosphide/zinc sulfide (InP/ZnS) QDs onto silica templates and coating them with a silica shell. The fabricated silica-coated InP/ZnS QD-embedded silica nanoparticles (SiO2@InP QDs@SiO2 NPs) exhibited hydrophilic properties because of the surface silica shell. The quantum yield (QY), maximum emission peak wavelength, and full-width half-maximum (FWHM) of the final fabricated SiO2@InP QDs@SiO2 NPs were 6.61%, 527.01 nm, and 44.62 nm, respectively. Moreover, the brightness of the particles could be easily controlled by adjusting the amount of InP/ZnS QDs in the SiO2@InP QDs@SiO2 NPs. When SiO2@InP QDs@SiO2 NPs were administered to tumor syngeneic mice, the fluorescence signal was prominently detected in the tumor because of the preferential distribution of the SiO2@InP QDs@SiO2 NPs, demonstrating their applicability in bioimaging with NPs. Thus, SiO2@InP QDs@SiO2 NPs have the potential to successfully replace Cd-based QDs as highly bright and biocompatible fluorescent nanoprobes.

Synthesis of Gold-Platinum Core-Shell Nanoparticles Assembled on a Silica Template and Their Peroxidase Nanozyme Properties.

Bimetallic nanoparticles are important materials for synthesizing multifunctional nanozymes. A technique for preparing gold-platinum nanoparticles (NPs) on a silica core template (SiO2@Au@Pt) using seed-mediated growth is reported in this study. The SiO2@Au@Pt exhibits peroxidase-like nanozyme activity has several advantages over gold assembled silica core templates (SiO2@Au@Au), such as stability and catalytic performance. The maximum reaction velocity (Vmax) and the Michaelis-Menten constants (Km) were and 2.1 × 10-10 M-1∙s-1 and 417 µM, respectively. Factors affecting the peroxidase activity, including the quantity of NPs, solution pH, reaction time, and concentration of tetramethyl benzidine, are also investigated in this study. The optimization of SiO2@Au@Pt NPs for H2O2 detection obtained in 0.5 mM TMB; using 5 µg SiO2@Au@Pt, at pH 4.0 for 15 min incubation. H2O2 can be detected in the dynamic liner range of 1.0 to 100 mM with the detection limit of 1.0 mM. This study presents a novel method for controlling the properties of bimetallic NPs assembled on a silica template and increases the understanding of the activity and potential applications of highly efficient multifunctional NP-based nanozymes.

Aptamer-conjugated nano-liposome for immunogenic chemotherapy with reversal of immunosuppression.

Cancer cells have various immune evasion mechanisms that resist the immune cells by reprogramming the tumor microenvironment (TME), such as programmed death-ligand 1 (PD-L1) and indoleamine 2,3-dioxygenase-1 (IDO1) overexpression. One of the approaches to restore antitumor immune response by T-cells is through induction of immunogenic cell death (ICD). Thus, drug carrier containing IDO1 siRNA and ICD inducer would be effective anticancer regimen to modulate the immunosuppressive TME by reversing the IDO1-mediated immunosuppression in a synergistic combination with ICD induction. However, numerous nanocarrier platforms for co-delivery of multiple drugs mostly depend on the enhanced permeation and retention (EPR), which is insufficient to achieve selectivity in tumor sites harboring various types of cells. We designed a targeted drug delivery system using nano-sized liposomes functionalized with anti-CD44 and anti-PD-L1 DNA aptamers, which target breast cancer cells and inhibit PD-1/PD-L1 interaction between cancer cells and T-cells. To reverse immunosuppressive TME and reactivate immune response, cancer-targeting nano-liposomes were prepared to contain immunogenic cell death inducer (Doxorubicin, DOX) and IDO1 siRNA, namely Aptm[DOX/IDO1]. The Aptm[DOX/IDO1] specifically delivered the loaded DOX and IDO1 siRNA into target breast cancer cells through aptamer-mediated endocytosis. Cancer-targeted DOX/IDO1 siRNA delivery enhanced ICD and suppressed IDO1 expression with significantly high toxicity in cancer cells. We demonstrated that Aptm[DOX/IDO1] could achieve synergistic antitumor effects by facilitating ICD response and simultaneous reversal of the immunosuppressive TME with IDO1 knockdown in the subcutaneous breast cancer model mice, thus reducing tumor size. These antitumor effects were exerted with intratumoral infiltration of CD8+ cytotoxic T lymphocyte as well as attenuation of regulatory T-cell recruitment in the tumor sites. We further proved that our Aptm[DOX/IDO1] strategy significantly reduced tumor metastasis in tumor-xenograft mice through a synergistic combination of cancer cell-targeted ICD induction and reversal of the IDO1-mediated immunosuppressive TME. Our nanocarrier platform based on cationic liposomes containing DOX and IDO1 siRNA, which are conjugated with two DNA aptamers targeting the cancer cell surface, accomplished synergistic chemoimmunotherapy through tumor-specific immune modulation into immune-favorable TME in vivo.

Highly specific chimeric DNA-RNA-guided genome editing with enhanced CRISPR-Cas12a system.

The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a system is composed of a Cas12a effector that acts as a DNA-cleaving endonuclease and a crispr RNA (crRNA) that guides the effector to the target DNA. It is considered a key molecule for inducing target-specific gene editing in various living systems. Here, we improved the efficiency and specificity of the CRISPR-Cas12a system through protein and crRNA engineering. In particular, to optimize the CRISPR-Cas12a system at the molecular level, we used a chimeric DNA-RNA guide chemically similar to crRNA to maximize target sequence specificity. Compared with the wild-type (wt)-Cas12a system, when using enhanced Cas12a system (en-Cas12a), the efficiency and target specificity improved on average by 2.58 and 2.77 times, respectively. In our study, when the chimeric DNA-RNA-guided en-Cas12a effector was used, the gene-editing efficiency and accuracy were simultaneously increased. These findings could contribute to highly accurate genome editing, such as human gene therapy, in the near future.

Expansion of the prime editing modality with Cas9 from Francisella novicida.

Prime editing can induce a desired base substitution, insertion, or deletion in a target gene using reverse transcriptase after nick formation by CRISPR nickase. In this study, we develop a technology that can be used to insert or replace external bases in the target DNA sequence by linking reverse transcriptase to the Francisella novicida Cas9, which is a CRISPR-Cas9 ortholog. Using FnCas9(H969A) nickase, the targeting limitation of existing Streptococcus pyogenes Cas9 nickase [SpCas9(H840A)]-based prime editing is dramatically extended, and accurate prime editing is induced specifically for the target genes in human cell lines.

Highly sensitive near-infrared SERS nanoprobes for in vivo imaging using gold-assembled silica nanoparticles with controllable nanogaps.

BACKGROUND: To take advantages, such as multiplex capacity, non-photobleaching property, and high sensitivity, of surface-enhanced Raman scattering (SERS)-based in vivo imaging, development of highly enhanced SERS nanoprobes in near-infrared (NIR) region is needed. A well-controlled morphology and biocompatibility are essential features of NIR SERS nanoprobes. Gold (Au)-assembled nanostructures with controllable nanogaps with highly enhanced SERS signals within multiple hotspots could be a breakthrough. RESULTS: Au-assembled silica (SiO2) nanoparticles (NPs) (SiO2@Au@Au NPs) as NIR SERS nanoprobes are synthesized using the seed-mediated growth method. SiO2@Au@Au NPs using six different sizes of Au NPs (SiO2@Au@Au50-SiO2@Au@Au500) were prepared by controlling the concentration of Au precursor in the growth step. The nanogaps between Au NPs on the SiO2 surface could be controlled from 4.16 to 0.98 nm by adjusting the concentration of Au precursor (hence increasing Au NP sizes), which resulted in the formation of effective SERS hotspots. SiO2@Au@Au500 NPs with a 0.98-nm gap showed a high SERS enhancement factor of approximately 3.8 × 106 under 785-nm photoexcitation. SiO2@Au@Au500 nanoprobes showed detectable in vivo SERS signals at a concentration of 16 µg/mL in animal tissue specimen at a depth of 7 mm. SiO2@Au@Au500 NPs with 14 different Raman label compounds exhibited distinct SERS signals upon subcutaneous injection into nude mice. CONCLUSIONS: SiO2@Au@Au NPs showed high potential for in vivo applications as multiplex nanoprobes with high SERS sensitivity in the NIR region.

Movable Layer Device for Rapid Detection of Influenza a H1N1 Virus Using Highly Bright Multi-Quantum Dot-Embedded Particles and Magnetic Beads.

Preventing the rapid spread of viral infectious diseases has become a major concern for global health. In this study, we present a microfluidic platform that performs an immunoassay of viral antigens in a simple, automated, yet highly sensitive manner. The device uses silica particles embedded with highly bright quantum dots (QD2) and performs the immunoassay with a vertically movable top layer and a rotating bottom layer. Through the motion of the layers and the surface tension in the liquids, reagents move from top chambers to bottom chambers and mix homogeneously. A tip in the top layer with a mobile permanent magnet moves the immune complexes comprising the magnetic beads, virus particles, and QD2 between the bottom chambers. In this way, our automated device achieves a highly sensitive magnetic bead-based sandwich immunoassay for the influenza A H1N1 virus within 32.5 min. The detection limit of our method is 5.1 × 10-4 hemagglutination units, which is 2 × 103 times more sensitive than that of the conventional hemagglutination method and is comparable to PCR. Our device is useful for the rapid and sensitive detection of infectious diseases in point-of-care applications and resource-limited environments.