Search

We developed a novel surface‐enhanced Raman scattering (SERS)‐based SARS‐CoV‐2 assay platform using hollow Au nanostars to realize high‐sensitivity diagnosis of SARS‐CoV‐2. The assay was performed using SARS‐CoV‐2 lysate as the target in a wide dynamic range with virus concentrations ranging from 0 to 104 PFU/ml and has a limit of detection (LOD) of 5.1 PFU/ml. This LOD value shows 100 times and 10 times better sensitivity compared to the LODs measured on the same sample using a commercially available rapid kit and enzyme‐linked immunosorbent assay, respectively. Therefore, we believe that this SERS‐based SARS‐CoV‐2 assay platform has high diagnostic accuracy for early or asymptomatic infected patients with low virus concentrations. Furthermore, the probability of a false‐negative diagnosis is likely to be very low., A new magnetic bead‐based SERS assay platform was developed to overcome the sensitivity limit of the commercially available SARS‐CoV‐2 immunodiagnostic kits. A novel hollow gold nanostars (HAuNSs), with a high enhancement factor and good stability, was employed for the detection of nucleocapsid protein from SARS‐CoV‐2. The assay could be performed in a wide dynamic range with virus concentrations ranging from 10−2 to 104 PFU/ml and showed an LOD value of 5.1 PFU/ml using this platform.

Raman thermometry based on surface-enhanced Raman scattering has been developed using nanopipettes in cancer cell photothermal therapy (PTT). Gold nanorods (AuNRs) are robustly epoxied on glass pipettes with a high surface coverage of ∼95% and less than 10 nm-wide nanogaps for intracellular thermometry and photothermal cancer therapy. The temperature changes could be estimated from the N≡C band shifts of 4-fluorophenyl isocyanide (FPNC)-adsorbed AuNRs on the Raman thermometry nanopipette (RTN) surfaces. An intracellular temperature change of ∼2.7 °C produced by altering the [Ca

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 β-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.

Enhanced carbon capture and oxygen production via water splitting was observed by controlling the plasmon-induced resonance energy transfer (PIRET) for photosystem II (PSII) in thylakoid extracts and spirulina assembled on gold nanoparticle (AuNP) dimer arrays. The two types of vertical (V) and horizontal (H) AuNP dimer arrays were uniformly inserted inside pore diameter-controlled templates. Based on the theoretical calculations, the longitudinal mode of the H AuNP dimer array was found to be sensitive to the nanogap distances between the two AuNPs in resonance with the absorption at P680 of the PSII. The longitudinal modes that interacted with P680 of PSII increased from the V to the H conformer. The optical properties from the H AuNP dimer array caused overlapping absorbance and photoluminescence with PSII, and the H AuNP dimer arrays exhibited a significant increase in carbon capture and oxygen generation rates in comparison with those of the bare PSII protein complex under light irradiation via the controlled PIRET process.

For the last two decades, surface-enhanced Raman spectroscopy (SERS) has been a useful bioanalytical tool for rapid and sensitive detection of biomolecules because of its highly sensitive and multiplex-detecting capability. In particular, SERS-based in vitro diagnostics combined with microfluidics offers target specificity, automatic sampling, and reduced use of analyte volumes. SERS is a highly sensitive detection modality, and microfluidics provides many advantages such as automatic sample handling and reduced sample volume. Therefore the integration of SERS with microfluidics offers extensive applications in biomedical diagnostics. In this chapter, recent advances in a wide range of SERS-based microfluidic devices such as continuous flow-based microfluidics, paper-based lateral flow assay strips, nano-patterned microarray-embedded microfluidics, and droplet-based microfluidics and their applications for in vitro diagnostics will be introduced.

A blue-, yellow- and red-emitting fluorescence sensor was constructed via the synergy of manganese dioxide (MnO2) nanosheets, o-phenylenediamine (OPD) and red-fluorescent quantum dots (r-QDs), namely MnO2–OPD–QD, for visualization of ascorbic acid (AA) and alkaline phosphatase (ALP). Initially, by virtue of the oxidase-like activity of MnO2 nanosheets, OPD was oxidized into yellow-fluorescent OPDox (emission: 569 nm), which quenched the r-QDs (emission: 659 nm). After interaction with AA, one reaction product of ALP and 2-phospho-L-ascorbic acid (AA2P), the MnO2 nanosheets were reduced and lost the oxidase-like activity, so that the generation of OPDox was lessened and the fluorescence of the r-QDs was thereupon recovered. Further increasing AA or ALP, the MnO2 nanosheets were fully decomposed so that OPD was not oxidized but was reduced into blue-fluorescent OPDred (emission: 441 nm), while the r-QDs were not affected. Overall, AA or ALP triggered emission intensity changes at 441, 569 and 659 nm and led to profuse color variation over the yellow–orange–red–purple range, realizing multiplex and visual determination of AA and ALP in the ranges of 0.5–70 μM and 0.1–100 mU mL−1, respectively. The color variation range of the ternary-emission mode was greatly enlarged, especially compared with that of the single-/dual-emission mode. Accordingly, the MnO2–OPD–QD sensor provided detection limits for AA and ALP as low as 0.18 μM and 0.06 mU mL−1, respectively. Furthermore, the highly selective/sensitive measurement and accurate readout of AA and ALP were accomplished in complicated food and serum samples. The multi-emitting fluorescence sensor held great potential for onsite visualization of various targets in more fields.

Surface-enhanced Raman scattering (SERS)-based assays have been recently developed to overcome the low detection sensitivity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SERS-based assays using magnetic beads in microtubes slightly improved the limit of detection (LoD) for SARS-CoV-2. However, the sensitivity and reproducibility of the method are still insufficient for reliable SARS-CoV-2 detection. In this study, we developed a SERS-based microdroplet sensor to dramatically improve the LoD and reproducibility of SARS-CoV-2 detection. Raman signals were measured for SERS nanotags in 140 droplets passing through a laser focal volume fixed at the center of the channel for 15 s. A comparison of the Raman signals of SERS nanotags measured in a microtube with those measured for multiple droplets in the microfluidic channel revealed that the LoD and coefficient of variation significantly improved from 36 to 0.22 PFU/mL and 21.2% to 1.79%, respectively. This improvement resulted from the ensemble average effects because the signals were measured for SERS nanotags in multiple droplets. Moreover, the total assay time decreased from 30 to 10 min. A clinical test was performed on patient samples to evaluate the clinical efficacy of the SERS-based microdroplet sensor. The assay results agreed well with those measured by the reverse transcription-polymerase chain reaction (RT-PCR) method. The proposed SERS-based microdroplet sensor is expected to be used as a new point-of-care diagnostic platform for quick and accurate detection of SARS-CoV-2 in the field.

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.

The surface-enhanced Raman scattering (SERS)-based lateral flow assay (LFA) strip has been considered a high-sensitivity sensor that can overcome the low sensitivity and the difficulty of quantitative analysis problems inherent in the colorimetric LFA sensor. In the SERS-based LFA strip reported so far, a liquid sample flows through the nitrocellulose membrane in a single pathway. In some cases, however, this single-flow approach still has a limitation in detection sensitivity. This study developed a conceptually new SERS-based dual-flow LFA sensor to improve the detection sensitivity in a single-flow LFA sensor. First, a 25 nm Raman reporter-labeled gold nanoparticle (AuNP) solution flowed through one way, and a 45 nm AuNP solution continuously flowed through another path. This sequential flow of two different AuNP solutions enables forming additional bright hot spots between 25 and 45 nm AuNPs in the test line, and the SERS signal is strongly enhanced. Using this SERS-based dual-flow LFA sensor, it was possible to detect thyroid-stimulating hormone less than 0.5 μIU/mL that cannot be measured with a SERS-based single-flow LFA sensor.

We introduce a facile and low-cost method for fabricating gold nanostructures on cellulose filter paper (CFP) to prepare a paper-based surface-enhanced Raman scattering (SERS) sensor for label-free molecular detection. Polymerized dopamine (PD) was used as an adhesive layer on the CFP and simultaneously functioned as a reducing agent for gold nanoparticle (AuNP) nucleation. The size of the AuNPs was dependent on the pH of the gold precursor solution, and nanoparticles with an average size of 102 nm were formed on the PD-coated CFP at a pH 3, exhibiting high SERS activity. Finite-difference time-domain (FDTD) simulations of the electromagnetic field enhancement of AuNPs with different sizes and interparticle distances were performed to identify the origin of the SERS effect. The developed paper-based SERS substrate showed uniform and excellent molecular sensitivity with a limit of detection (LOD) of 10−7 M for methylene blue, as measured by a portable Raman spectrometer. Furthermore, as a field application test, surfaces of apples were pretreated with diquat (DQ) and paraquat (PQ) pesticides, which were then detected down to a concentration of 1 ppm after simple attachment of the sensor on the apple peels and performing a SERS measurement. The developed paper-based SERS sensor is expected to be applicable as a label-free sensor for a variety of chemical and biological molecules.

We have visualized surface-enhanced Raman scattering (SERS) nano-gap domains with electrostatic force microscopy (EFM). We constructed the SERS substrate using reactive-ion etching of a polyethylene terephthalate (PET) film, with subsequent sputtering of Au onto the etched PET. Because the nano-gap is more strongly charged than the native Au-nanoparticle (NP) surface, we can identify the SERS nano-gap due to the longer dwell time of the tapping EFM tip and the related phase shift of the signal. This produces a differentiating image contrast between the SERS nano-gap and the Au NP surface, with a spatial resolution of ~14 nm. More specifically, differences in the mode-independent Raman-signal enhancement and the peak shift of an adsorbed malachite green isothiocyanate (MGITC) molecule are only observed in the smaller SERS nano-gap (found in the surface with Ar+-sputtered Au nano-pillars). We interpret this in terms of the differing morphologies of the underlying Au substrates and the related mechanical compressive strains, which result in charge transfer (CT)-based single-molecule Raman-reporter detection sensitivity.

Green ion imprinted polymers (IIPs) were prepared in aqueous phase via the synergy of three functional monomers of low-cost eco-friendly gelatin (G), 8-hydroxyquinoline (HQ) and chitosan (C), namely G-HQ-C IIPs, and were applied as an effective and recyclable adsorbent to remove Cu(II) from aqueous solution. The as-prepared G-HQ-C IIPs were systematically characterized, and several major factors affecting adsorption capacity including solution pH, temperature and contact time were investigated in detail. The adsorption of Cu(II) on G-HQ-C IIPs followed the pseudo-second-order kinetic and Langmuir isotherm models, and the adsorption capacity increased with temperature increase. Moreover, the maximum adsorption capacities of G-HQ-C IIPs toward Cu(II) reached up to 111.81 mg/g at room temperature, much higher than those of most of the reported adsorbents for Cu(II). The G-HQ-C IIPs displayed excellent selectivity against seven common divalent ions with selectivity coefficients above 18.71, as well as high anti-interference ability. Additionally, a good reusability was demonstrated without significant loss in adsorption capacity after at least ten cycles. The IIPs were applied to environmental water samples for selective removal of Cu(II) with satisfactory results. By replacing Cu(II) template by Cd(II), Hg(II) and Pb(II), respectively, the obtained three kinds of IIPs based on G-HQ-C presented convincing imprinting properties, and therefore the work could provide a simple and general imprinting strategy toward various concerned heavy metal ions through multi-point interactions from multiple functional monomers.

Recently, microfluidic-based flow cytometry platforms have been shown to be powerful tools for the manipulation and analysis of single cells and micron-sized particles in flow. That said, current microfluidic flow cytometers are limited in both their analytical throughput and spatial resolution, due to their reliance on single point interrogation schemes. Conversely, high-speed imaging techniques can be applied to a wide variety of problems in which analyte molecules are manipulated at high linear velocities. Such an approach allows a detailed visualization of dynamic events through acquisition of a series of image frames captured with high temporal and spatial resolution. Herein, we describe some of the most significant recent advances in the development of multi-parametric, optofluidic imaging flow cytometry for the enumeration of complex cellular populations.

Theranostics nanoplatforms offer opportunities for imaging-guided precision therapy and hold great potential for clinical applications. In most reported works, the imaging unit has a lack of site selectivity, and is always kept in the "on" modality regardless of whether it is in normal tissues or tumor sites, increasing the risk of unsafe treatment. Herein, we designed a near-infrared (NIR) fluorescence-guided theranostics nanoplatform by integrating the functions of tumor-response and photodynamic therapy (PDT)/photothermal therapy (PTT). A novel NIR fluorescent dye, CyPT, with excellent optical and PDT/PTT properties, was synthesized and linked onto the gold nanorods (AuNRs) to form CyPT-AuNRs nanohybrids via a sulfur-sulfur bond that can be broken by glutathione (GSH) with high selectivity and sensitivity. In normal cells where the concentration of GSH is low, the fluorescence of CyPT is quenched by the AuNRs. By contrast, the high level of GSH in tumor cells leads to the breaking of the sulfur-sulfur bond, resulting in the release of CyPT and the accomplishment of a "off-on" fluorescence response. Followed by precise NIR tumor-imaging diagnosis, the PDT and PTT treatment which rely on the released CyPT and AuNRs, respectively, can be effectively performed. The CyPT-AuNRs nanoplatform has been successfully applied to the treatment of tumor xenograft models and no distinct damage has been observed in the nearby normal tissues. This versatile nanoplatform has potential for use in targeted tumor imaging and precision therapy.

The phage display technique is a combinatorial technology in which random peptides are displayed on the surface of the phage; it is widely used to identify high-affinity peptides that bind to a target protein. However, this technique presents several problems due to non-specific binding of the phages and steric hindrance caused by blocking agents. To overcome these problems, we tested two modified methods and compared their screening performance with that of the conventional method. We used poly-His-tagged human epidermal growth factor receptor 2 (HER2) as a target protein and silica-coated magnetic particles (MPs) with an immobilized nickel-nitrilotriacetic acid ligand as a solid matrix. Modified method #1 (#M1) included two negative selection steps against a blocking agent (bovine serum albumin) and nude Ni-NTA MPs, after the positive selection step using immobilized HER2 on MPs in the absence of BSA. Modified method #2 (#M2) allowed the binding of phages and HER2 in solution prior to immobilizing HER2 on the MP surface. The negative selection procedure was the same between them. The binding affinity of the phages screened by #M1 and #M2 was evaluated by phage ELISA. Two phages from #M2 (#M2-4 and 5) showed the highest binding, and between them #M2-5 was selected for affinity maturation by inserting a hairpin-structured peptide between the two #M2-5 peptides. SPR study showed the HER2-binding affinity was indeed improved by ca. 2000-fold to become comparable with that of the intact antibody (trastuzumab). The modified biopanning methods combined with the dimerization maturation can be an effective way to develop antibody-mimicking peptides for affinity binding.

In the last two decades, microfluidic technology has emerged as a highly efficient tool for the study of various chemical and biological reactions. Recently, we reported that high-throughput detection of various concentrations of a reagent is possible using a continuous gradient microfluidic channel combined with a surface-enhanced Raman scattering (SERS) detection platform. In this continuous flow regime, however, the deposition of nanoparticle aggregates on channel surfaces induces the "memory effect," affecting both sensitivity and reproducibility. To resolve this problem, a SERS-based gradient droplet system was developed. Herein, the serial dilution of a reagent was achieved in a stepwise manner using microfluidic concentration gradient generators. Then various concentrations of a reagent generated in different channels were simultaneously trapped into the tiny volume of droplets by injecting an oil mixture into the channel. Compared to the single-phase regime, this two-phase liquid/liquid segmented flow regime allows minimization of resident time distributions of reagents through localization of reagents in encapsulated droplets. Consequently, the sample stacking problem could be solved using this system because it greatly reduces the memory effect. We believe that this SERS-based gradient droplet system will be of significant utility in simultaneously monitoring chemical and biological reactions for various concentrations of a reagent.

The use of fluorescent probes in visible detection has been developed over the last several decades. Biomolecules are essential in the biological processes of organisms, and their distribution and concentration are largely influenced by environmental factors. Significant advances have occurred in the applications of fluorescent probes for the detection of the dynamic localization and quantity of biomolecules during various environmental stress-induced physiological and pathological processes. Herein, we summarize representative examples of small molecule-based fluorescent probes that provide bimolecular information when the organism is under environmental stress. The discussion includes strategies for the design of smart small-molecule fluorescent probes, in addition to their applications in biomolecule imaging under environmental stresses, such as hypoxia, ischemia-reperfusion, hyperthermia/hypothermia, organic/inorganic chemical exposure, oxidative/reductive stress, high glucose stimulation, and drug treatment-induced toxicity. We believe that comprehensive insight into the beneficial applications of fluorescent probes in biomolecule detection under environmental stress should enable the further development and effective application of fluorescent probes in the biochemical and biomedical fields.

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.

Bacterial pathogens such as Yersinia pestis, Francisella tularensis, and Bacillus anthracis are classified into the highest rank of potential bioterrorism agents. Colorimetric lateral flow assay (LFA) strips are commercially available but these conventional strips have drawbacks in terms of low sensitivity and limit of quantitative analysis. Therefore, there is an urgent need for a new sensing platform to detect these pathogens in the early contamination stage. In this study, a novel surface-enhanced Raman scattering (SERS)-based LFA strip was developed for sensitive detection of bacterial pathogens. Target-specific SERS nanotags (Raman reporter-labeled gold nanoparticles) were used as an alternative to the gold nanoparticles in conventional LFA strips. Using these SERS nanotags the presence of bacteria could be identified through a simple color change in the test line. Additionally, highly sensitive and accurate quantitative analysis could be performed by monitoring the characteristic Raman peak intensity of SERS nanotags that were captured in the test line. This highly sensitive method required a short assay time (15 min) and a tiny volume of pathogen sample (40 μL). We believe that the proposed SERS-based LFA technique has great potential as a valuable tool in the early detection of specific bacterial pathogens in the field due to its excellent analytical sensitivity.

Advances in revolutionary technologies pose new challenges for human life; in response to them, global responsibility is pushing modern technologies toward greener pathways. Molecular imprinting technology (MIT) is a multidisciplinary mimic technology simulating the specific binding principle of enzymes to substrates or antigens to antibodies; along with its rapid progress and wide applications, MIT faces the challenge of complying with green sustainable development requirements. With the identification of environmental risks associated with unsustainable MIT, a new aspect of MIT, termed green MIT, has emerged and developed. However, so far, no clear definition has been provided to appraise green MIT. Herein, the implementation process of green chemistry in MIT is demonstrated and a mnemonic device in the form of an acronym, GREENIFICATION, is proposed to present the green MIT principles. The entire greenificated imprinting process is surveyed, including element choice, polymerization implementation, energy input, imprinting strategies, waste treatment, and recovery, as well as the impacts of these processes on operator health and the environment. Moreover, assistance of upgraded instrumentation in deploying greener goals is considered. Finally, future perspectives are presented to provide a more complete picture of the greenificated MIT road map and to pave the way for further development.

This minireview reports the recent advances in surface-enhanced Raman scattering (SERS)-based assay devices for the diagnosis of infectious diseases. SERS-based detection methods have shown promise in overcoming the low sensitivity and multiplex detection problems inherent to fluorescence detection. Therefore, it is interesting to investigate the current status, challenges, and applications associated with SERS-based microdevices for the point-of-care (POC) diagnosis of infectious diseases. The majority of this review highlights three different types of microdevices, namely microfluidic channels, lateral flow assay strips, and three-dimensional nanostructured substrates. Furthermore, the integration of portable Raman spectrophotometry with microdevices provides an ideal platform for the diagnosis of various infectious diseases in the field. Integrated SERS-based assay systems also enable measurements in minimal sample volumes and at low analyte concentrations of viral or bacterial samples. A significant number of studies using the SERS-based assay system have been performed recently to realize POC diagnostics, especially under resource-limited conditions. This portable SERS sensor is expected to be a next-generation POC assay system that could overcome the limitations of current fluorescence-based assay systems. This minireview summarizes recent advances in the development of SERS-based microdevices for the diagnosis of infectious diseases. Lastly, challenges to overcome and future perspectives are discussed.

Test strips represent a class of point-of-care testing (POCT) tools for analysis of a variety of biomarkers towards diagnostics. Conventional test strips offer benefits of simple operation, visualization, and short detection time, along with the drawbacks of relatively low sensitivity and unavailability of quantitative analysis. Recently, the combination of surface-enhanced Raman scattering (SERS) and test strips have evolved to provide a powerful platform capable of ultrasensitive and multiplex detection of extensive analytes of interest. In this review, we focus on the working principles, design strategies and POCT applications of SERS-based test strips. Initially, both lateral and vertical flow test strips are briefly introduced, followed by presentation of various strategies for reforming SERS-based test strips with better detection performance. Applications of SERS-based test strips in diagnosis of disease biomarkers, nucleic acids and toxins are reviewed, with an emphasis on SERS tag design, sensitivity and analytical applicability. Finally, conclusions are made and perspectives on futuristic research directions are given.

We developed a dual-mode surface-enhanced Raman scattering (SERS)-based aptasensor that can accurately diagnose and distinguish severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A/H1N1 at the same time. Herein, DNA aptamers that selectively bind to SARS-CoV-2 and influenza A/H1N1 were immobilized together on Au nanopopcorn substrate. Raman reporters (Cy3 and RRX), attached to the terminal of DNA aptamers, could generate strong SERS signals in the nanogap of the Au nanopopcorn substrate. Additionally, the internal standard Raman reporter (4-MBA) was immobilized on the Au nanopopcorn substrate along with aptamer DNAs to reduce errors caused by changes in the measurement environment. When SARS-CoV-2 or influenza A virus approaches the Au nanopopcorn substrate, the corresponding DNA aptamer selectively detaches from the substrate due to the significant binding affinity between the corresponding DNA aptamer and the virus. As a result, the related SERS intensity decreases with increasing target virus concentration. Thus, it is possible to determine whether a suspected patient is infected with SARS-CoV-2 or influenza A using this SERS-based DNA aptasensor. Furthermore, this sensor enables a quantitative evaluation of the target virus concentration with high sensitivity without being affected by cross-reactivity. Therefore, this SERS-based diagnostic platform is considered a conceptually new diagnostic tool that rapidly discriminates against these two respiratory diseases to prevent their spread.

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.

Hypoxic stress is a common concern in medicine and biology, which can induce the cellular injury and death by excess production of reactive oxygen species (ROS). Hypochlorous acid (HOCl), one of the ROS, plays a crucial role in the oxidative damage to tissue proteins in the pathogenesis of various diseases. Therefore, excess production of HOCl might be an important factor for the damage caused by hypoxic stress. However, most of the present methods cannot meet the demand of real-time detection on account of the labile and reactive chemical properties of HOCl. Herein, we designed a near-infrared fluorescent probe, Cy-HOCl, for the selective imaging of HOCl in cells and in vivo. Cy-HOCl includes two moieties: a 4-amino-3-nitrophenol group as the response unit and a near-infrared heptamethine cyanine fluorophore as the fluorescent modulator. Cy-HOCl exhibits excellent selectivity and sensitivity towards the detection of HOCl. The hypoxic response behavior of Cy-HOCl is evaluated in cells to clarify the relationship between HOCl and hypoxia. The probe is also applied to measure HOCl in ex vivo-dissected organs of an acute ischemia mouse model as well as for real-time monitoring the changes of HOCl in the hypoxic zebrafish model.

We report the analysis of deoxyribonuclease (DNase) activity by conjugation-free fluorescence polarisation in a droplet-based microfluidic chip. DNase is a DNA cleaving enzyme and its activity is important in the maintenance of normal cellular functions. Alterations in DNase activity have been implicated as the cause of various cancers and autoimmune diseases. To date, various methods for the analysis of DNase activity have been reported. However, they are not cost effective due to the requirement of large sample volumes and the need for the conjugation of fluorescent dyes. In this study, we have used ethidium bromide (EtBr), a DNA intercalating reagent, as a fluorescent reporter without any prior conjugation or modification of DNA. Degradation of DNA by DNase 1 was monitored at a steady state by making changes in the fluorescence polarisation of EtBr in droplets with a volume of 330 picolitre at a 40 hertz frequency under visible light. Using this technique, we successfully determined the half-maximal inhibitory concentration (IC50) of ethylenediaminetetraacetic acid (EDTA) for the inhibition of DNase 1 activity to be 1.56 ± 0.91 mM.

There is a challenge to obtain an ultrasensitive and rapid approach to detect bacteria and identify resistance. As a powerful bioanalytical tool, surface-enhanced Raman scattering (SERS) in bacterial detection have attracted increasing attentions. Herein, we developed a SERS-active Au@Ag core-shell nanorod (Au@AgNR) tag platform for ultrasensitive bacteria detection and antibiotic-susceptibility testing (AST). The platform established that surface enhanced Raman scattered Rhodamine 6G (R6G) absorption at 1517 cm−1 had a good linearity (RI = 3865 + 193logC; R2 = 0.97) with logarithm of E. coli concentration over a range of 107–102 CFU (colony forming unit)/mL with limit of detection as low 102 CFU/mL. When E. coli was exposed to ampicillin at minimum inhibitory concentration (MIC, 4 μg/mL), Raman spectroscopy showed the obvious variation between ampicillin-susceptible E. coli (Amp−-E. coli) and the ampicillin-resistant E. coli (Amp+-E. coli). Combined with principal component analysis (PCA) statistical analysis, the Raman intensity variation mentioned above allows to obtain rapid antibiotic resistance testing (

The design and fabrication of multifunctional surface-enhanced Raman scattering (SERS) nanotags are key issues in their application to biological imaging of cells and tissues. In this study, highly sensitive, reproducible and long-term stable SERS nanotags were developed for the identification of localized distribution of multiple protein biomarkers expressed on breast cancer cells. To enhance the surface electromagnetic fields of Raman reporter molecules, Ag-encapsulated Au (Ag-Au) hollow nanospheres were synthesized. Strong Raman signal enhancement effects could be achieved by positioning Raman reporter molecules in nanogaps between the Au hollow nanospheres and silver shell. In addition, the signal was also enhanced due to the localization of surface electromagnetic fields through the pinholes on the surface of Au hollow nanospheres. To maintain the long-term stability of the Au hollow-Ag core/shell nanospheres, their surface was coated with a polyethylene glycol (PEG) layer. The biocompatibility of PEGylated Ag-Au hollow nanospheres was investigated using the premix water soluble tetrazolium salt (WST-1) cell viability test. These SERS nanotags also enabled a high-resolution multiplexed live cell imaging. Our proposed SERS imaging technique using the new SERS nanotags provides a new platform for fast and accurate classification of different phenotypes of breast cancer cells.

To date, 55% of inhabitants of the developing world still live in rural regions and they have a very urgent need to improve the level of disease diagnosis, especially in resource-limited settings. Herein, we present a microfluidic system that centrifuges whole blood and quantifies contained biomarkers in a rapid, feasible and integrated way. Such a device provides a blood-in/answer-out capability and is small enough to be carried by any individual in any environment. The successful integration a hand-powered centrifuge and immunoassay unit within a rotational paper-based device allows for diagnostic application by untrained users and in environments where access to electricity cannot be assumed. In addition, the low cost (~$ 0.5), light weight and small instrumental footprint make the device ideally suited for rapid on-site detection. To validate the applicability of the system in a clinical diagnostic testing, we successfully perform enzyme-linked immunosorbent assay (ELISA) analysis of carcinoembryonic antigen and alpha fetoprotein from human blood samples. We expect that this powerful platform technology will provide the opportunities for point-of-care diagnosis in resource-limited settings.

We report a surface-enhanced Raman scattering (SERS)-based polymerase chain reaction (PCR) assay platform for the sensitive and rapid detection of a DNA marker (pagA) of Bacillus anthracis. Real-ti...

A variety of automated sample-in-answer-out systems for in vitro molecular diagnostics have been presented and even commercialized. Although efficient in operation, they are incapable of quantifying targets, since quantitation based on analog analytical methods (via standard curve analysis) is complex, expensive, and challenging. To address this issue, herein, we describe an integrated sample-in-digital-answer-out (SIDAO) diagnostic system incorporating DNA extraction and digital recombinase polymerase amplification, which enables rapid and quantitative nucleic acid analysis from bodily fluids within a disposable cartridge. Inside the cartridge, reagents are pre-stored in sterilized tubes, with an automated pipetting module allowing facile liquid transfer. For digital analysis, we fabricate a simple, single-layer polydimethylsiloxane microfluidic device and develop a novel and simple sample compartmentalization strategy. Sample solution is partitioned into an array of 40,044 fL-volume microwells by sealing the microfluidic device through the application of mechanical pressure. The entire analysis is performed in a portable, fully automated instrument. We evaluate the quantitative capabilities of the system by analyzing Mycobacterium tuberculosis genomic DNA from both spiked saliva and serum samples, and demonstrate excellent analytical accuracy and specificity. This SIDAO system provides a promising diagnostic platform for quantitative nucleic acid testing at the point-of-care. Graphical abstract ᅟ.

Flexible tactile sensors have been intensively studied for healthcare and electronic skin devices. Currently, a sensing material, electrode, and substrate are manufactured as one set by depositing the sensing material on the electrode. For this reason, when another electrode or substrate is used in the sensor or when different sensor characteristics are required, a new sensing material must be developed and the fabrication conditions should be changed. This study proposes a novel method of manufacturing a pressure sensing material like a cartridge film. The cartridge film is made by filling the holes of a stencil film (one MP in each hole) with conductive microparticles (MPs). Using the cartridge film, the sensing material can be cut-and-pasted on electrodes and transferred to other electrodes for reuse. This study analyzes the electrical responses of the sensors made of the cartridge film on the basis of the Hertzian contact theory, and also correlates the sensing performance of the sensors with the cond...

In recent years there has been huge interest in the development of microfluidic reactors for the synthesis of small molecules and nanomaterials. Such reaction platforms represent a powerful and versatile alternative to traditional formats since they allow for the precise, controlled, and flexible management of reactive processes. To date, the majority of microfluidic reactors used in small-molecule synthesis have been manufactured using conventional lithographic techniques from materials such as glasses, ceramics, stainless steel, and silicon. Surprisingly, the fabrication of microfluidic devices from such rigid materials remains ill-defined, complex, and expensive. Accordingly, the microfluidic toolkit for chemical synthesis would significantly benefit from the development of solvent-resistant microfluidic devices that can be manufactured using soft-lithographic prototyping methods. Whilst significant advances in the development of solvent-resistant polymers have been made, only modest steps have been taken towards simplifying their use as microfluidic reactors. Herein, we emphasize the benefits of using a commercially available, amorphous perfluorinated polymer, CYTOP, as a coating with which to transform PDMS into a chemically inert material for use in organic synthesis applications. Its efficacy is demonstrated through the subsequent performance of photooxidation reactions and reactions under extremely acidic or basic conditions.

We utilized a fast Raman spectral mapping technique for fast detection of bacterial pathogens. Three-dimensional (3D) plasmonic nanopillar arrays were fabricated using the nanolithography-free process consisting of maskless Ar plasma treatment of a polyethylene terephthalate substrate and subsequent metal deposition. Bacterial pathogens were immobilized on the positively charged poly(l-lysine)-coated 3D plasmonic substrate through electrostatic interactions. Then, the bacterial surfaces were selectively labeled with antibody-conjugated surface-enhanced Raman scattering (SERS) nanotags, and Raman mapping images were collected and statistically analyzed for quantitative analysis of bacteria. Salmonella typhimurium was selected as a model pathogen bacterium to confirm the efficacy of our SERS imaging technique. Minimum number of Raman mapping points with statistical reliability was determined to reduce assay time. It was possible to get a statistically reliable standard calibration curve for 529 pixels (laser spot with 60 μm interval), which required a total mapping time of 45 min to get a standard calibration curve for five different concentrations of bacteria in the 0 to 10