Search results for: plasmon resonace

Authors: Emineh Tsegahun Gedif

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Zinc oxide nanoparticles (ZnO NPs) have garnered significant attention due to their diverse applications encompassing catalytic, optical, photonic, and antibacterial properties. In this study, we successfully synthesized zinc oxide nanoparticles using a rapid, environmentally benign, and cost-effective method. Neem (Azadirachta indica) leaf extract served as the reducing agent for Zn (NO₃)₂.6H2O solution under optimized conditions (pH = 9). Qualitative screening techniques and FT-IR Spectroscopy confirmed the presence of active biomolecules such as flavonoids, phenolic groups, alkaloids, terpenoids, and tannins within the Neem leaf extract, both before and after reduction. The formation of ZnO NPs was visually evident through a distinct color change from colorless to light yellow. The biosynthesized nanoparticles underwent comprehensive characterization through UV-visible, FT-IR, and XRD spectroscopies. The reduction process proved to be straightforward and user-friendly, with UV-visible spectroscopy demonstrating a surface plasmon resonance (SPR) at 321 nm, unequivocally confirming the ZnO NP formation. X-ray diffraction analysis elucidated the crystal structure, revealing an average particle size of approximately 20 nm using Scherrer's equation based on the line width of the plane. Furthermore, the synthesized zinc oxide nanoparticles were evaluated for their antimicrobial properties against both Gram-positive and Gram-negative bacteria. The results showcased significant inhibitory activity, with the highest zone of inhibition observed against Escherichia coli (15 mm) and comparatively lower activity against Staphylococcus aureus. This research underscores the potential of Neem leaf extract-mediated synthesis of ZnO NPs as an eco-friendly and effective approach for various applications, including antibacterial agents.

Keywords: zinc oxide nanoparticles (ZnO NPs), bioreducing agent, green synthesis, antibacterial activity

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Authors: Christian Fercher, Jiaul Islam, Simon R. Corrie

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Fluorescence-based immunodiagnostics are an emerging field in biosensor development and exhibit several advantages over traditional detection methods. While various affinity biosensors have been developed to generate a fluorescence signal upon sensing varying concentrations of analytes, reagentless, reversible, and continuous monitoring of complex biological samples remains challenging. Here, we aimed to genetically engineer biosensors based on single-chain antibody fragments (scFv) that are site-specifically labeled with environmentally sensitive fluorescent unnatural amino acids (UAA). A rational design approach resulted in quantifiable analyte-dependent changes in peak fluorescence emission wavelength and enabled antigen detection in vitro. Incorporation of a polarity indicator within the topological neighborhood of the antigen-binding interface generated a titratable wavelength blueshift with nanomolar detection limits. In order to ensure continuous analyte monitoring, scFv candidates with fast binding and dissociation kinetics were selected from a genetic library employing a high-throughput phage display and affinity screening approach. Initial rankings were further refined towards rapid dissociation kinetics using bio-layer interferometry (BLI) and surface plasmon resonance (SPR). The most promising candidates were expressed, purified to homogeneity, and tested for their potential to detect biomarkers in a continuous microfluidic-based assay. Variations of dissociation kinetics within an order of magnitude were achieved without compromising the specificity of the antibody fragments. This approach is generally applicable to numerous antibody/antigen combinations and currently awaits integration in a wide range of assay platforms for one-step protein quantification.

Keywords: antibody engineering, biosensor, phage display, unnatural amino acids

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Authors: Mohammed Alasel, Michael Keusgen

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Gold is a noble metal; in its nano-scale level (e.g. spherical nanoparticles), the conduction electrons are triggered to collectively oscillate with a resonant frequency when certain wavelengths of electromagnetic radiation interact with its surface; this phenomenon is known as surface plasmon resonance (SPR). SPR is responsible for giving the gold nanoparticles its intense red color depending mainly on its size, shape and distance between nanoparticles. A decreased distance between gold nanoparticles results in aggregation of them causing a change in color from red to blue. This aggregation enables gold nanoparticles to serve as a sensitive biosensoric indicator. In the proposed work, gold nanoparticles were modified with two proteins: i) Borrelia antigen, variable lipoprotein surface-exposed protein (VlsE), and ii) protein A. VlsE antigen induces a strong antibody response against Lyme disease and can be detected from early to late phase during the disease in humans infected with Borrelia. In addition, it shows low cross-reaction with the other non-pathogenic Borrelia strains. The high specificity of VlsE antigen to anti-Borrelia antibodies, combined simultaneously with the high specificity of protein A to the Fc region of all IgG human antibodies, was utilized to develop a rapid test for serological point of care diagnosis of borreliosis in human serum. Only in the presence of anti-Borrelia antibodies in the serum probe, an aggregation of gold nanoparticles can be observed, which is visible by a concentration-dependent colour shift from red (low IgG) to blue (high IgG). Experiments showed it is clearly possible to distinguish between positive and negative sera samples using a simple suspension of the two-protein modified gold nanoparticles in a very short time (30 minutes). The proposed work showed the potential of using such modified gold nanoparticles generally for serological diagnosis. Improved specificity and reduced assay time can be archived in applying increased salt concentrations combined with decreased pH values (pH 5).

Keywords: gold nanoparticles, gold aggregation, serological diagnosis, protein A, lyme borreliosis

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Authors: Lucja Rodzik, Joanna Lewandowska-Lancucka, Michal Szuwarzynski, Krzysztof Szczubialka, Maria Nowakowska

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The analysis of nucleobases is of great importance for bioscience since their abnormal concentration in body fluids suggests the deficiency and mutation of the immune system, and it is considered to be an important parameter for diagnosis of various diseases. The presented conjugate meets the need for development of the effective, selective and highly sensitive sensor for nucleobase/nucleoside detection. The novel, highly fluorescent cadmium telluride quantum dots (CdTe QDs) functionalized with thymine and stabilized with thioglycolic acid (TGA) conjugates has been developed and thoroughly characterized. Successful formation of the material was confirmed by elemental analysis, and UV–Vis fluorescence and FTIR spectroscopies. The crystalline structure of the obtained product was characterized with X-ray diffraction (XRD) method. The composition of CdTe QDs and their thymine conjugate was also examined using X-ray photoelectron spectroscopy (XPS). The size of the CdTe-thymine was 3-6 nm as demonstrated using atomic force microscopy (AFM) and high resolution transmission electron microscopy (HRTEM) imaging. The plasmon resonance fluorescence band at 540 nm on excitation at 351 nm was observed for these nanoparticles. The intensity of this band increased with the increase in the amount of conjugated thymine with no shift in its position. Based on the fluorescence measurements, it was found that the CdTe-thymine conjugate interacted efficiently and selectively not only with adenine, a nucleobase complementary to thymine, but also with nucleosides and adenine-containing modified nucleosides, i.e., 5′-deoxy-5′-(methylthio)adenosine (MTA) and 2’-O-methyladenosine, the urinary tumor markers which allow monitoring of the disease progression. The applicability of the CdTe-thymine sensor for the real sample analysis was also investigated in simulated urine conditions. High sensitivity and selectivity of CdTe-thymine fluorescence towards adenine, adenosine and modified adenosine suggest that obtained conjugate can be potentially useful for development of the biosensor for complementary nucleobase/nucleoside detection.

Keywords: CdTe quantum dots, conjugate, sensor, thymine

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Authors: Elmira Solati, Atousa Mehrani, Davoud Dorranian

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Generation of ZnO-Au nanocomposite under laser irradiation of a mixture of the ZnO and Au colloidal suspensions are experimentally investigated. In this work, firstly ZnO and Au nanoparticles are prepared by pulsed laser ablation of the corresponding metals in water using the 1064 nm wavelength of Nd:YAG laser. In a second step, the produced ZnO and Au colloidal suspensions were mixed in different volumetric ratio and irradiated using the second harmonic of a Nd:YAG laser operating at 532 nm wavelength. The changes in the size of the nanostructure and optical properties of the ZnO-Au nanocomposite are studied as a function of the volumetric ratio of ZnO and Au colloidal suspensions. The crystalline structure of the ZnO-Au nanocomposites was analyzed by X-ray diffraction (XRD). The optical properties of the samples were examined at room temperature by a UV-Vis-NIR absorption spectrophotometer. Transmission electron microscopy (TEM) was done by placing a drop of the concentrated suspension on a carbon-coated copper grid. To further confirm the morphology of ZnO-Au nanocomposites, we performed Scanning electron microscopy (SEM) analysis. Room temperature photoluminescence (PL) of the ZnO-Au nanocomposites was measured to characterize the luminescence properties of the ZnO-Au nanocomposites. The ZnO-Au nanocomposites were characterized by Fourier transform infrared (FTIR) spectroscopy. The X-ray diffraction pattern shows that the ZnO-Au nanocomposites had the polycrystalline structure of Au. The behavior observed by images of transmission electron microscope reveals that soldering of Au and ZnO nanoparticles include their adhesion. The plasmon peak in ZnO-Au nanocomposites was red-shifted and broadened in comparison with pure Au nanoparticles. By using the Tauc’s equation, the band gap energy for ZnO-Au nanocomposites is calculated to be 3.15–3.27 eV. In this work, the formation of ZnO-Au nanocomposites shifts the FTIR peak of metal oxide bands to higher wavenumbers. PL spectra of the ZnO-Au nanocomposites show that several weak peaks in the ultraviolet region and several relatively strong peaks in the visible region. SEM image indicates that the morphology of ZnO-Au nanocomposites produced in water was spherical. The TEM images of ZnO-Au nanocomposites demonstrate that with increasing the volumetric ratio of Au colloidal suspension the adhesion increased. According to the size distribution graphs of ZnO-Au nanocomposites with increasing the volumetric ratio of Au colloidal suspension the amount of ZnO-Au nanocomposites with the smaller size is further.

Keywords: Au nanoparticles, pulsed laser ablation, ZnO-Au nanocomposites, ZnO nanoparticles

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Authors: Siamak Dawazdah Emami, Amin Khodaei, Harith Bin Ahmad, Hairul A. Adbul-Rashid

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The Fano resonance effect on plasmonic nanoparticle materials results in such materials possessing a number of unique optical properties, and the potential applicability for sensing, nonlinear devices and slow-light devices. A Fano resonance is a consequence of coherent interference between superradiant and subradiant hybridized plasmon modes. Incident light on subradiant modes will initiate excitation that results in superradiant modes, and these superradient modes possess zero or finite dipole moments alongside a comparable negligible coupling with light. This research work details the derivation of an electrodynamics coupling model for the interaction of dipolar transitions and radiation via plasmonic nanoclusters such as quadrimers, pentamers and heptamers. The directivity calculation is analyzed in order to qualify the redirection of emission. The geometry of a configured array of nanostructures strongly influenced the transmission and reflection properties, which subsequently resulted in the directivity of each antenna being related to the nanosphere size and gap distances between the nanospheres in each model’s structure. A well-separated configuration of nanospheres resulted in the structure behaving similarly to monomers, with spectra peaks of a broad superradiant mode being centered within the vicinity of 560 nm wavelength. Reducing the distance between ring nanospheres in pentamers and heptamers to 20~60 nm caused the coupling factor and charge distributions to increase and invoke a subradiant mode centered within the vicinity of 690 nm. Increasing the outside ring’s nanosphere distance from the centered nanospheres caused the coupling factor to decrease, with the coupling factor being inversely proportional to cubic of the distance between nanospheres. This phenomenon led to a dramatic decrease of the superradiant mode at a 200 nm distance between the central nanosphere and outer rings. Effects from a superradiant mode vanished beyond a 240 nm distance between central and outer ring nanospheres.

Keywords: fano resonance, optical antenna, plasmonic, nano-clusters

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Authors: Pavel Damborsky, Martina Zamorova, Jaroslav Katrlik

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Prostate cancer is annually the most common newly diagnosed cancer among men. An extensive number of evidence suggests that traditional serum Prostate-specific antigen (PSA) assay still suffers from a lack of sufficient specificity and sensitivity resulting in vast over-diagnosis and overtreatment. Thus, the early-stage detection of prostate cancer (PCa) plays undisputedly a critical role for successful treatment and improved quality of life. Over the last decade, particular altered glycans have been described that are associated with a range of chronic diseases, including cancer and inflammation. These glycans differences enable a distinction to be made between physiological and pathological state and suggest a valuable biosensing tool for diagnosis and follow-up purposes. Aberrant glycosylation is one of the major characteristics of disease progression. Consequently, the aim of this study was to develop a more reliable tool for early-stage PCa diagnosis employing lectins as glyco-recognition elements. Biosensor and biochip technology putting to use lectin-based glyco-profiling is one of the most promising strategies aimed at providing fast and efficient analysis of glycoproteins. The proof-of-concept experiments based on sandwich assay employing anti-PSA antibody and an aptamer as a capture molecules followed by lectin glycoprofiling were performed. We present a lectin-based biosensing assay for glycoprofiling of serum biomarker PSA using different biosensor and biochip platforms such as label-free surface plasmon resonance (SPR) and microarray with fluorescent label. The results suggest significant differences in interaction of particular lectins with PSA. The antibody-based assay is frequently associated with the sensitivity, reproducibility, and cross-reactivity issues. Aptamers provide remarkable advantages over antibodies due to the nucleic acid origin, stability and no glycosylation. All these data are further step for construction of highly selective, sensitive and reliable sensors for early-stage diagnosis. The experimental set-up also holds promise for the development of comparable assays with other glycosylated disease biomarkers.

Keywords: biomarker, glycosylation, lectin, prostate cancer

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Authors: Hana Hanaee-Ahvaz, Monika Cserjan-Puschmann, Christopher Tauer, Gerald Striedner

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Incorporation of noncanonical amino acids (ncAA) into proteins has become an interesting topic as proteins featured with ncAAs offer a wide range of different applications. Nowadays, technologies and systems exist that allow for the site-specific introduction of ncAAs in vivo, but the efficient production of proteins modified this way is still a big challenge. This is especially true for 'hard-to-express' proteins where low yields are encountered even with the native sequence. In this study, site-specific incorporation of azido-ethoxy-carbonyl-Lysin (azk) into an anti-tumor-necrosis-factor-α-Fab (FTN2) was investigated. According to well-established parameters, possible site positions for ncAA incorporation were determined, and corresponding FTN2 genes were constructed. Each of the modified FTN2 variants has one amber codon for azk incorporated either in its heavy or light chain. The expression level for all variants produced was determined by ELISA, and all azk variants could be produced with a satisfactory yield in the range of 50-70% of the original FTN2 variant. In terms of expression yield, neither the azk incorporation position nor the subunit modified (heavy or light chain) had a significant effect. We confirmed correct protein processing and azk incorporation by mass spectrometry analysis, and antigen-antibody interaction was determined by surface plasmon resonance analysis. The next step is to characterize the effect of azk incorporation on protein stability and aggregation tendency via differential scanning calorimetry and light scattering, respectively. In summary, the incorporation of ncAA into our Fab candidate FTN2 worked better than expected. The quantities produced allowed a detailed characterization of the variants in terms of their properties, and we can now turn our attention to potential applications. By using click chemistry, we can equip the Fabs with additional functionalities and make them suitable for a wide range of applications. We will now use this option in a first approach and develop an assay that will allow us to follow the degradation of the recombinant target protein in vivo. Special focus will be laid on the proteolytic activity in the periplasm and how it is influenced by cultivation/induction conditions.

Keywords: degradation, FTN2, hard-to-express protein, non-canonical amino acids

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Authors: Yunha Ryu, Kyoungsik Kim

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Raman spectroscopy is one of the most powerful techniques for chemical detection, but the low sensitivity originated from the extremely small cross-section of the Raman scattering limits the practical use of Raman spectroscopy. To overcome this problem, Surface Enhanced Raman Scattering (SERS) has been intensively studied for several decades. Because the SERS effect is mainly induced from strong electromagnetic near-field enhancement as a result of localized surface plasmon resonance of metallic nanostructures, it is important to design the plasmonic structures with high density of electromagnetic hot spots for SERS substrate. One of the useful fabrication methods is using porous nanomaterial as a template for metallic structure. Internal pores on a scale of tens of nanometers can be strong EM hotspots by confining the incident light. Also, porous structures can capture more target molecules than non-porous structures in a same detection spot thanks to the large surface area. Herein we report the facile fabrication method of porous SERS substrate by integrating solvent-assisted nanoimprint lithography and selective etching of block copolymer. We obtained nanostructures with high porosity via simple selective etching of the one microdomain of the diblock copolymer. Furthermore, we imprinted of the nanocone patterns into the spin-coated flat block copolymer film to make three-dimensional SERS substrate for the high density of SERS hot spots as well as large surface area. We used solvent-assisted nanoimprint lithography (SAIL) to reduce the fabrication time and cost for patterning BCP film by taking advantage of a solvent which dissolves both polystyrenre and poly(methyl methacrylate) domain of the block copolymer, and thus block copolymer film was molded under the low temperature and atmospheric pressure in a short time. After Ag deposition, we measured Raman intensity of dye molecules adsorbed on the fabricated structure. Compared to the Raman signals of Ag coated solid nanocone, porous nanocone showed 10 times higher Raman intensity at 1510 cm(-1) band. In conclusion, we fabricated porous metallic nanocone arrays with high density electromagnetic hotspots by templating nanoimprinted diblock copolymer with selective etching and demonstrated its capability as an effective SERS substrate.

Keywords: block copolymer, porous nanostructure, solvent-assisted nanoimprint, surface-enhanced Raman spectroscopy

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Authors: Masauso Moses Phiri

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Frequent glucose monitoring is essential to the management of diabetes. Plasmonic enzyme-based glucose biosensors have the advantages of greater specificity, simplicity and rapidity. The aim of this study was to develop a rapid plasmonic colorimetric glucose biosensor based on biocatalytic enlargement of AuNS guided by GOx. Gold nanoparticles of 18 nm in diameter were synthesized using the citrate method. Using these as seeds, a modified seeded method for the synthesis of monodispersed gold nanostars was followed. Both the spherical and star-shaped nanoparticles were characterized using ultra-violet visible spectroscopy, agarose gel electrophoresis, dynamic light scattering, high-resolution transmission electron microscopy and energy-dispersive X-ray spectroscopy. The feasibility of a plasmonic colorimetric assay through growth of AuNS by silver coating in the presence of hydrogen peroxide was investigated by several control and optimization experiments. Conditions for excellent sensing such as the concentration of the detection solution in the presence of 20 µL AuNS, 10 mM of 2-(N-morpholino) ethanesulfonic acid (MES), ammonia and hydrogen peroxide were optimized. Using the optimized conditions, the glucose assay was developed by adding 5mM of GOx to the solution and varying concentrations of glucose to it. Kinetic readings, as well as color changes, were observed. The results showed that the absorbance values of the AuNS were blue shifting and increasing as the concentration of glucose was elevated. Control experiments indicated no growth of AuNS in the absence of GOx, glucose or molecular O₂. Increased glucose concentration led to an enhanced growth of AuNS. The detection of glucose was also done by naked-eye. The color development was near complete in ± 10 minutes. The kinetic readings which were monitored at 450 and 560 nm showed that the assay could discriminate between different concentrations of glucose by ± 50 seconds and near complete at ± 120 seconds. A calibration curve for the qualitative measurement of glucose was derived. The magnitude of wavelength shifts and absorbance values increased concomitantly with glucose concentrations until 90 µg/mL. Beyond that, it leveled off. The lowest amount of glucose that could produce a blue shift in the localized surface plasmon resonance (LSPR) absorption maxima was found to be 10 – 90 µg/mL. The limit of detection was 0.12 µg/mL. This enabled the construction of a direct sensitivity plasmonic colorimetric detection of glucose using AuNS that was rapid, sensitive and cost-effective with naked-eye detection. It has great potential for transfer of technology for point-of-care devices.

Keywords: colorimetric, gold nanostars, glucose, glucose oxidase, plasmonic

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Authors: Piya Roychoudhury, Ruma Pal

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Cyanobacteria, commonly known as blue green algae were found to be an effective bioreagent for nanoparticle synthesis. Nowadays silver nanoparticles (AgNPs) are very popular due to their antimicrobial and anti-proliferative activity. To exploit these characters in different biotechnological fields, it is very essential to synthesize more stable, non-toxic nano-silver. For this reason silver-gold alloy (Ag-AuNPs) nanoparticles are of great interest as they are more stable, harder and more effective than single metal nanoparticles. In the present communication we described a simple technique for rapid synthesis of biocompatible AgNP and Ag-AuNP employing cyanobacteria, Leptolyngbya and Lyngbya respectively. For synthesis of AgNP the biomass of Leptolyngbya valderiana (200 mg Fresh weight) was exposed to 9 mM AgNO3 solution (pH 4). For synthesis of Ag-AuNP Lyngbya majuscula (200 mg Fresh weight) was exposed to equimolar solution of hydrogen tetra-auro chlorate and silver nitrate (1mM, pH 4). After 72 hrs of exposure thallus of Leptolyngyba turned brown in color and filaments of Lyngbya turned pink in color that indicated synthesis of nanoparticles. The produced particles were extracted from the cyanobacterial biomass using nano-capping agent, sodium citrate. Firstly, extracted brown and pink suspensions were taken for Energy Dispersive X-ray (EDAX) analysis to confirm the presence of silver in brown suspension and presence of both gold and silver in pink suspension. Extracted nanoparticles showed a distinct single plasmon band (AgNP at 411 nm; Ag-Au NP at 481 nm) in Uv-vis spectroscopy. It was revealed from Transmission electron microscopy (TEM) that all the synthesized particles were spherical in nature with a size range of ~2-25 nm. In X-ray powder diffraction (XRD) analysis four intense peaks appeared at 38.2°, 44.5°, 64.8°and 77.8° which confirmed the crystallographic nature of synthesized particles. Presence of different functional groups viz. N-H, C=C, C–O, C=O on the surface of nanoparticles were recorded by Fourier transform infrared spectroscopy (FTIR). Scanning Electron microscopy (SEM) images showed the surface topography of metal treated filaments of cyanobacteria. The stability of the particles was observed by Zeta potential study. Antibiotic property of synthesized particles was tested by Agar well diffusion method against gram negative bacteria Pseudomonas aeruginosa. Overall, this green-technique requires low energy, less manufacturing cost and produces rapidly eco-friendly metal nanoparticles.

Keywords: cyanobacteria, silver nanoparticles, silver-gold alloy nanoparticles, spectroscopy

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Authors: Sulalit Bandyopadhyay, Muhammad Awais Ashfaq Alvi, Anuvansh Sharma, Wilhelm R. Glomm

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Release of drugs from nanogels and nanogel-based systems can occur under the influence of external stimuli like temperature, pH, magnetic fields and so on. pNIPAm-AAc nanogels respond to the combined action of both temperature and pH, the former being mostly determined by hydrophilic-to-hydrophobic transitions above the volume phase transition temperature (VPTT), while the latter is controlled by the degree of protonation of the carboxylic acid groups. These nanogels based systems are promising candidates in the field of drug delivery. Combining nanogels with magneto-plasmonic nanoparticles (NPs) introduce imaging and targeting modalities along with stimuli-response in one hybrid system, thereby incorporating multifunctionality. Fe@Au core-shell NPs possess optical signature in the visible spectrum owing to localized surface plasmon resonance (LSPR) of the Au shell, and superparamagnetic properties stemming from the Fe core. Although there exist several synthesis methods to control the size and physico-chemical properties of pNIPAm-AAc nanogels, yet, there is no comprehensive study that highlights the dependence of incorporation of one or more layers of NPs to these nanogels. In addition, effective determination of volume phase transition temperature (VPTT) of the nanogels is a challenge which complicates their uses in biological applications. Here, we have modified the swelling-collapse properties of pNIPAm-AAc nanogels, by combining with Fe@Au NPs using different solution based methods. The hydrophilic-hydrophobic transition of the nanogels above the VPTT has been confirmed to be reversible. Further, an analytical method has been developed to deduce the average VPTT which is found to be 37.3°C for the nanogels and 39.3°C for nanogel coated Fe@Au NPs. An opposite swelling –collapse behaviour is observed for the latter where the Fe@Au NPs act as bridge molecules pulling together the gelling units. Thereafter, Cyt C, a model protein drug and L-Dopa, a drug used in the clinical treatment of Parkinson’s disease were loaded separately into the nanogels and nanogel coated Fe@Au NPs, using a modified breathing-in mechanism. This gave high loading and encapsulation efficiencies (L Dopa: ~9% and 70µg/mg of nanogels, Cyt C: ~30% and 10µg/mg of nanogels respectively for both the drugs. The release kinetics of L-Dopa, monitored using UV-vis spectrophotometry was observed to be rather slow (over several hours) with highest release happening under a combination of high temperature (above VPTT) and acidic conditions. However, the release of L-Dopa from nanogel coated Fe@Au NPs was the fastest, accounting for release of almost 87% of the initially loaded drug in ~30 hours. The chemical structure of the drug, drug incorporation method, location of the drug and presence of Fe@Au NPs largely alter the drug release mechanism and the kinetics of these nanogels and Fe@Au NPs coated with nanogels.

Keywords: controlled release, nanogels, volume phase transition temperature, l-dopa

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Authors: Moustafa Osman Mohammed

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This paper introduces topological order in descried social systems starting with the original concept of autopoiesis by biologists and scientists, including the modification of general systems based on socialized medicine. Topological order is important in describing the physical systems for exploiting optical systems and improving photonic devices. The stats of topological order have some interesting properties of topological degeneracy and fractional statistics that reveal the entanglement origin of topological order, etc. Topological ideas in photonics form exciting developments in solid-state materials, that being; insulating in the bulk, conducting electricity on their surface without dissipation or back-scattering, even in the presence of large impurities. A specific type of autopoiesis system is interrelated to the main categories amongst existing groups of the ecological phenomena interaction social and medical sciences. The hypothesis, nevertheless, has a nonlinear interaction with its natural environment 'interactional cycle' for exchange photon energy with molecules without changes in topology. The engineering topology of a biosensor is based on the excitation boundary of surface electromagnetic waves in photonic band gap multilayer films. The device operation is similar to surface Plasmonic biosensors in which a photonic band gap film replaces metal film as the medium when surface electromagnetic waves are excited. The use of photonic band gap film offers sharper surface wave resonance leading to the potential of greatly enhanced sensitivity. So, the properties of the photonic band gap material are engineered to operate a sensor at any wavelength and conduct a surface wave resonance that ranges up to 470 nm. The wavelength is not generally accessible with surface Plasmon sensing. Lastly, the photonic band gap films have robust mechanical functions that offer new substrates for surface chemistry to understand the molecular design structure and create sensing chips surface with different concentrations of DNA sequences in the solution to observe and track the surface mode resonance under the influences of processes that take place in the spectroscopic environment. These processes led to the development of several advanced analytical technologies: which are; automated, real-time, reliable, reproducible, and cost-effective. This results in faster and more accurate monitoring and detection of biomolecules on refractive index sensing, antibody-antigen reactions with a DNA or protein binding. Ultimately, the controversial aspect of molecular frictional properties is adjusted to each other in order to form unique spatial structure and dynamics of biological molecules for providing the environment mutual contribution in investigation of changes due to the pathogenic archival architecture of cell clusters.

Keywords: autopoiesis, photonics systems, quantum topology, molecular structure, biosensing

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Authors: Ewelina Grabowska, Martyna Marchelek

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Among the various semiconductors, nanosized TiO2 has been widely studied due to its high photosensitivity, low cost, low toxicity, and good chemical and thermal stability. However, there are two main drawbacks to the practical application of pure TiO2 films. One is that TiO2 can be induced only by ultraviolet (UV) light due to its intrinsic wide bandgap (3.2 eV for anatase and 3.0 eV for rutile), which limits its practical efficiency for solar energy utilization since UV light makes up only 4-5% of the solar spectrum. The other is that a high electron-hole recombination rate will reduce the photoelectric conversion efficiency of TiO2. In order to overcome the above drawbacks and modify the electronic structure of TiO2, some semiconductors (eg. CdS, ZnO, PbS, Cu2O, Bi2S3, and CdSe) have been used to prepare coupled TiO2 composites, for improving their charge separation efficiency and extending the photoresponse into the visible region. It has been proved that the fabrication of p-n heterostructures by combining n-type TiO2 with p-type semiconductors is an effective way to improve the photoelectric conversion efficiency of TiO2. SrTiO3 is a good candidate for coupling TiO2 and improving the photocatalytic performance of the photocatalyst because its conduction band edge is more negative than TiO2. Due to the potential differences between the band edges of these two semiconductors, the photogenerated electrons transfer from the conduction band of SrTiO3 to that of TiO2. Conversely, the photogenerated electrons transfer from the conduction band of SrTiO3 to that of TiO2. Then the photogenerated charge carriers can be efficiently separated by these processes, resulting in the enhancement of the photocatalytic property in the photocatalyst. Additionally, one of the methods for improving photocatalyst performance is addition of nanoparticles containing one or two noble metals (Pt, Au, Ag and Pd) deposited on semiconductor surface. The mechanisms were proposed as (1) the surface plasmon resonance of noble metal particles is excited by visible light, facilitating the excitation of the surface electron and interfacial electron transfer (2) some energy levels can be produced in the band gap of TiO2 by the dispersion of noble metal nanoparticles in the TiO2 matrix; (3) noble metal nanoparticles deposited on TiO2 act as electron traps, enhancing the electron–hole separation. In view of this, we recently obtained series of TiO2@SrTiO3 and SrTiO3 photocatalysts loaded with noble metal NPs. using photodeposition method. The M- TiO2@SrTiO3 and M-SrTiO3 photocatalysts (M= Rh, Rt, Pt) were studied for photodegradation of phenol in aqueous phase under UV-Vis and visible irradiation. Moreover, in the second part of our research hydroxyl radical formations were investigated. Fluorescence of irradiated coumarin solution was used as a method of ˙OH radical detection. Coumarin readily reacts with generated hydroxyl radicals forming hydroxycoumarins. Although the major hydroxylation product is 5-hydroxycoumarin, only 7-hydroxyproduct of coumarin hydroxylation emits fluorescent light. Thus, this method was used only for hydroxyl radical detection, but not for determining concentration of hydroxyl radicals.

Keywords: composites TiO2, SrTiO3, photocatalysis, phenol degradation

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Authors: Monisha Elumalai, Joana Guerreiro, Joana Carvalho, Marta Prado

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DNA biosensors popularity has been increasing over the past few years. Traditional analytical techniques tend to require complex steps and expensive equipment however DNA biosensors have the advantage of getting simple, fast and economic. Additionally, the combination of DNA biosensors with nanomaterials offers the opportunity to improve the selectivity, sensitivity and the overall performance of the devices. DNA biosensors are based on oligonucleotides as sensing elements. These oligonucleotides are highly specific to complementary DNA sequences resulting in the hybridization of the strands. DNA biosensors are not only an advantage in the clinical field but also applicable in numerous research areas such as food analysis or environmental control. Zebra Mussels (ZM), Dreissena polymorpha are invasive species responsible for enormous negative impacts on the environment and ecosystems. Generally, the detection of ZM is made when the observation of adult or macroscopic larvae's is made however at this stage is too late to avoid the harmful effects. Therefore, there is a need to develop an analytical tool for the early detection of ZM. Here, we present a portable plasmonic biosensor for the detection of environmental DNA (eDNA) released to the environment from this invasive species. The plasmonic DNA biosensor combines gold nanoparticles, as transducer elements, due to their great optical properties and high sensitivity. The detection strategy is based on the immobilization of a short base pair DNA sequence on the nanoparticles surface followed by specific hybridization in the presence of a complementary target DNA. The hybridization events are tracked by the optical response provided by the nanospheres and their surrounding environment. The identification of the DNA sequences (synthetic target and probes) to detect Zebra mussel were designed by using Geneious software in order to maximize the specificity. Moreover, to increase the optical response enzyme amplification of DNA might be used. The gold nanospheres were synthesized and characterized by UV-visible spectrophotometry and transmission electron microscopy (TEM). The obtained nanospheres present the maximum localized surface plasmon resonance (LSPR) peak position are found to be around 519 nm and a diameter of 17nm. The DNA probes modified with a sulfur group at one end of the sequence were then loaded on the gold nanospheres at different ionic strengths and DNA probe concentrations. The optimal DNA probe loading will be selected based on the stability of the optical signal followed by the hybridization study. Hybridization process leads to either nanoparticle dispersion or aggregation based on the presence or absence of the target DNA. Finally, this detection system will be integrated into an optical sensing platform. Considering that the developed device will be used in the field, it should fulfill the inexpensive and portability requirements. The sensing devices based on specific DNA detection holds great potential and can be exploited for sensing applications in-loco.

Keywords: ZM DNA, DNA probes, nicking enzyme, gold nanoparticles

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Authors: Sutapa Mondal Roy, Suban K. Sahoo

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The primary aim of this work is to synthesis silver nanoparticles (AgNPs) through environmentally benign routes to avoid any chemical toxicity related undesired side effects. The nanoparticles were stabilized with drug ciprofloxacin (Cp) and were studied for their effectiveness as drug delivery agent. Targeted drug delivery improves the therapeutic potential of drugs at the diseased site as well as lowers the overall dose and undesired side effects. The small size of nanoparticles greatly facilitates the transport of active agents (drugs) across biological membranes and allows them to pass through the smallest capillaries in the body that are 5-6 μm in diameter, and can minimize possible undesired side effects. AgNPs are non-toxic, inert, stable, and has a high binding capacity and thus can be considered as biomaterials. AgNPs were synthesized from the nutrient broth supernatant after the culture of environment-friendly bacteria Bacillus subtilis. The AgNPs were found to show the surface plasmon resonance (SPR) band at 425 nm. The Cp capped Ag nanoparticles formation was complete within 30 minutes, which was confirmed from absorbance spectroscopy. Physico-chemical nature of the AgNPs-Cp system was confirmed by Dynamic Light Scattering (DLS), Transmission Electron Microscopy (TEM) etc. The AgNPs-Cp system size was found to be in the range of 30-40 nm. To monitor the kinetics of drug release from the surface of nanoparticles, the release of Cp was carried out by careful dialysis keeping AgNPs-Cp system inside the dialysis bag at pH 7.4 over time. The drug release was almost complete after 30 hrs. During the drug delivery process, to understand the AgNPs-Cp system in a better way, the sincere theoretical investigation is been performed employing Density Functional Theory. Electronic charge transfer, electron density, binding energy as well as thermodynamic properties like enthalpy, entropy, Gibbs free energy etc. has been predicted. The electronic and thermodynamic properties, governed by the AgNPs-Cp interactions, indicate that the formation of AgNPs-Cp system is exothermic i.e. thermodynamically favorable process. The binding energy and charge transfer analysis implies the optimum stability of the AgNPs-Cp system. Thus, the synthesized Cp-Ag nanoparticles can be effectively used for biological purposes due to its environmentally benign routes of synthesis procedures, which is clean, biocompatible, non-toxic, safe, cost-effective, sustainable and eco-friendly. The Cp-AgNPs as biomaterials can be successfully used for drug delivery procedures due to slow release of drug from nanoparticles over a considerable period of time. The kinetics of the drug release show that this drug-nanoparticle assembly can be effectively used as potential tools for therapeutic applications. The ease of synthetic procedure, lack of possible chemical toxicity and their biological activity along with excellent application as drug delivery agent will open up vista of using nanoparticles as effective and successful drug delivery agent to be used in modern days.

Keywords: silver nanoparticles, ciprofloxacin, density functional theory, drug delivery

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Authors: Swati Mishra

Abstract:

In recent years, there has been an explosion in Gold NanoParticle (GNP) research, with a rapid increase in publications in diverse fields, including imaging, bioengineering, and molecular biology. GNPs exhibit unique physicochemical properties, including surface plasmon resonance (SPR) and bind amine and thiol groups, allowing surface modification and use in biomedical applications. Nanoparticle functionalization is the subject of intense research at present, with rapid progress being made towards developing biocompatible, multi-functional particles. In the present study, the photochemical method has been done to functionalize various-shaped GNPs like nanostars by the molecules like ninhydrin. Ninhydrin is bactericidal, virucidal, fungicidal, antigen-antibody reactive, and used in fingerprint technology in forensics. The GNPs functionalized with ninhydrin efficiently will bind to the amino acids on the target protein, which is of eminent importance during the pandemic, especially where long-term treatments of COVID- 19 bring many side effects of the drugs. The photochemical method is adopted as it provides low thermal load, selective reactivity, selective activation, and controlled radiation in time, space, and energy. The GNPs exhibit their characteristic spectrum, but a distinctly blue or redshift in the peak will be observed after UV irradiation, ensuring efficient ninhydrin binding. Now, the bound ninhydrin in the GNP carrier, upon chemically reacting with any amino acid, will lead to the formation of Rhumann purple. A common method of GNP production includes citrate reduction of Au [III] derivatives such as aurochloric acid (HAuCl4) in water to Au [0] through a one-step synthesis of size-tunable GNPs. The following reagents are prepared to validate the approach. Reagent A solution 1 is0.0175 grams ninhydrin in 5 ml Millipore water Reagent B 30 µl of HAuCl₄.3H₂O in 3 ml of solution 1 Reagent C 1 µl of gold nanostars in 3 ml of solution 1 Reagent D 6 µl of cetrimonium bromide (CTAB) in 3 ml of solution1 ReagentE 1 µl of gold nanostars in 3 ml of ethanol ReagentF 30 µl of HAuCl₄.₃H₂O in 3 ml of ethanol ReagentG 30 µl of HAuCl₄.₃H₂O in 3 ml of solution 2 ReagentH solution 2 is0.0087 grams ninhydrin in 5 ml Millipore water ReagentI 30 µl of HAuCl₄.₃H₂O in 3 ml of water The reagents were irradiated at 254 nm for 15 minutes, followed by their UV Visible spectroscopy. The wavelength was selected based on the one reported for excitation of a similar molecule Pthalimide. It was observed that the solution B and G deviate around 600 nm, while C peaks distinctively at 567.25 nm and 983.9 nm. Though it is tough to say about the chemical reaction happening, butATR-FTIR of reagents will ensure that ninhydrin is not forming Rhumann purple in the absence of amino acids. Therefore, these experiments, we achieved the functionalization of gold nanostars with ninhydrin corroborated by the deviation in the spectrum obtained in a mixture of GNPs and ninhydrin irradiated with UV light. It prepares them as a carrier molecule totake up amino acids for targeted delivery or germicidal action.

Keywords: gold nanostars, ninhydrin, photochemical method, UV visible specgtroscopy

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Authors: Jessie Rapoza, Petr Moroshkin, Jimmy Xu

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Chirality is omnipresent, in nature, in life, and in the field of physics. One intriguing example is the homochirality that has remained a great secret of life. Another is the pairs of mirror-image molecules – enantiomers. They are identical in atomic composition and therefore indistinguishable in the scalar physical properties. Yet, they can be either therapeutic or toxic, depending on their chirality. Recent studies suggest a potential link between abnormal levels of certain D-amino acids and some serious health impairments, including schizophrenia, amyotrophic lateral sclerosis, and potentially cancer. Although indistinguishable in their scalar properties, the chirality of a molecule reveals itself in interaction with the surrounding of a certain chirality, or more generally, a broken mirror-symmetry. In this work, we report on a system for chiral molecule detection, in which the mirror-symmetry is doubly broken, first by asymmetric structuring a nanopatterned plasmonic surface than by the incidence of circularly polarized light (CPL). In this system, the incident circularly-polarized light induces a surface plasmon polariton (SPP) wave, propagating along the asymmetric plasmonic surface. This SPP field itself is chiral, evanescently bound to a near-field zone on the surface (~10nm thick), but with an amplitude greatly intensified (by up to 104) over that of the incident light. It hence probes just the molecules on the surface instead of those in the volume. In coupling to molecules along its path on the surface, the chiral SPP wave favors one chirality over the other, allowing for chirality detection via the change in an optical rectification current measured at the edges of the sample. The asymmetrically structured surface converts the high-frequency electron plasmonic-oscillations in the SPP wave into a net DC drift current that can be measured at the edge of the sample via the mechanism of optical rectification. The measured results validate these design concepts and principles. The observed optical rectification current exhibits a clear differentiation between a pair of enantiomers. Experiments were performed by focusing a 1064nm CW laser light at the sample - a gold grating microchip submerged in an approximately 1.82M solution of either L-arabinose or D-arabinose and water. A measurement of the current output was then recorded under both rights and left circularly polarized lights. Measurements were recorded at various angles of incidence to optimize the coupling between the spin-momentums of the incident light and that of the SPP, that is, spin-momentum locking. In order to suppress the background, the values of the photocurrent for the right CPL are subtracted from those for the left CPL. Comparison between the two arabinose enantiomers reveals a preferential signal response of one enantiomer to left CPL and the other enantiomer to right CPL. In sum, this work reports on the first experimental evidence of the feasibility of chiral molecule detection via optical rectification in a metal meta-grating. This nanoscale interfaced electrical detection technology is advantageous over other detection methods due to its size, cost, ease of use, and integration ability with read-out electronic circuits for data processing and interpretation.

Keywords: Chirality, detection, molecule, spin

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Authors: Yi Li, Rui Lu, Lianjun Wang

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With the rapid development of chemical industry, the consumption of volatile organic compounds (VOCs) has increased extensively. In the process of VOCs production and application, plenty of them have been transferred to environment. As a result, it has led to pollution problems not only in soil and ground water but also to human beings. Thus, it is important to develop a sensitive and cost-effective analytical method for trace VOCs detection in environment. Surface-enhanced Raman Spectroscopy (SERS), as one of the most sensitive optical analytical technique with rapid response, pinpoint accuracy and noninvasive detection, has been widely used for ultratrace analysis. Based on the plasmon resonance on the nanoscale metallic surface, SERS technology can even detect single molecule due to abundant nanogaps (i.e. 'hot spots') on the nanosubstrate. In this work, a self-supported flexible silver nitrate (AgNO3)/ethylene-propylene copolymer (EPM) hybrid nanofibers was fabricated by electrospinning. After an in-situ chemical reduction using ice-cold sodium borohydride as reduction agent, numerous silver nanoparticles were formed on the nanofiber surface. By adjusting the reduction time and AgNO3 content, the morphology and dimension of silver nanoparticles could be controlled. According to the principles of solid-phase extraction, the hydrophobic substance is more likely to partition into the hydrophobic EPM membrane in an aqueous environment while water and other polar components are excluded from the analytes. By the enrichment of EPM fibers, the number of hydrophobic molecules located on the 'hot spots' generated from criss-crossed nanofibers is greatly increased, which further enhances SERS signal intensity. The as-prepared Ag/EPM hybrid nanofibers were first employed to detect common SERS probe molecule (p-aminothiophenol) with the detection limit down to 10-12 M, which demonstrated an excellent SERS performance. To further study the application of the fabricated substrate for monitoring hydrophobic substance in water, several typical VOCs, such as benzene, toluene and p-xylene, were selected as model compounds. The results showed that the characteristic peaks of these target analytes in the mixed aqueous solution could be distinguished even at a concentration of 10-6 M after multi-peaks gaussian fitting process, including C-H bending (850 cm-1), C-C ring stretching (1581 cm-1, 1600 cm-1) of benzene, C-H bending (844 cm-1 ,1151 cm-1), C-C ring stretching (1001 cm-1), CH3 bending vibration (1377 cm-1) of toluene, C-H bending (829 cm-1), C-C stretching (1614 cm-1) of p-xylene. The SERS substrate has remarkable advantages which combine the enrichment capacity from EPM and the Raman enhancement of Ag nanoparticles. Meanwhile, the huge specific surface area resulted from electrospinning is benificial to increase the number of adsoption sites and promotes 'hot spots' formation. In summary, this work provides powerful potential in rapid, on-site and accurate detection of trace VOCs using a portable Raman.

Keywords: electrospinning, ethylene-propylene copolymer, silver nanoparticles, SERS, VOCs

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Authors: Maria Zulema Morquecho Vega, Fabiola CarolinaMiranda Castro, Rafael Verdugo Miranda, Ignacio Yocupicio Villegas, Ana lidia Barron Raygoza, Martin enrique MArquez Cordova, Jose Alberto Duarte Moller

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Background: Anemopsis californica, also called (tame grass) belongs to the Saururaceae family small, green plant. The blade is long and wide. Gives a white flower. The plant population is only found in humid, swampy habitats, it grows where there is water, along the banks of streams and water holes. In the winter, it dries up. The leaves, rhizomes, or roots of this plant have been used to treat a range of diseases. Some of its healing properties are used to treat wounds, cold and flu symptoms, spasmodic cough, infection, pain and inflammation, burns, swollen feet, as well as lung ailments, asthma, circulatory problems (varicose veins), rheumatoid arthritis, purifies blood, helps in urinary and digestive tract diseases, sores and healing, for headache, sore throat, diarrhea, kidney pain. The tea made from the leaves and roots is used to treat uterine cancer, womb cancer, relieves menstrual pain and stops excessive bleeding after childbirth. It is also used as a gynecological treatment for infections, hemorrhoids, candidiasis and vaginitis. Objective: To study the cytotoxicity of gels prepared with silver nanoparticles in AC extract combined with chitosan, collagen and hyaluronic acid as an alternative therapy for skin conditions. Methods: The Ag NPs were synthesized according to the following method. A 0.3 mg/mL solution is prepared in 10 ml of deionized water, adjust to pH 12 with NaOH, stirring is maintained constant magnetic and a temperature of 80 °C. Subsequently, 100 ul of a 0.1 M AgNO3 solution and kept stirring constantly for 15 min. Once the reaction is complete, measurements are performed by UV-Vis. A gel was prepared in a 5% solution of acetic acid with the respective nanoparticles and AC extract of silver in the extract of AC. Chitosan is added until the process begins to occur gel. At that time, collagen will be added in a ratio of 3 to 5 drops, and later, hyaluronic acid in 2% of the total compound formed. Finally, after resting for 24 hours, the cytotoxic effect of the gels was studied. in the presence of highly positive bacteria Staphylococcus aureus and highly negative for Escherichia coli. Cultures will be incubated for 24 hours in the presence of the compound and compared with the reference. Results: Silver nanoparticles obtained had a spherical shape and sizes among 20 and 30 nm. UV-Vis spectra confirm the presence of silver nanoparticles showing a surface plasmon around 420 nm. Finally, the test in presence of bacteria yield a good antibacterial property of this nanocompound and tests in people were successful. Conclusion: Gel prepared by biogenic synthesis shown beneficious effects in severe acne, acne vulgaris and wound healing with diabetic patients.

Keywords: anemopsis californica, nanomedicina, biotechnology, biomedicine

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Authors: Francesca Crivellari, Nicholas Mavrogiannis, Zachary Gagnon

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Portable diagnostic methods have become increasingly important for a number of different purposes: point-of-care screening in developing nations, environmental contamination studies, bio/chemical warfare agent detection, and end-user use for commercial health monitoring. The cheapest and most portable methods currently available are paper-based – lateral flow and dipstick methods are widely available in drug stores for use in pregnancy detection and blood glucose monitoring. These tests are successful because they are cheap to produce, easy to use, and require minimally invasive sampling. While adequate for their intended uses, in the realm of blood-borne pathogens and numerous cancers, these paper-based methods become unreliable, as they lack the nM/pM sensitivity currently achieved by clinical diagnostic methods. Clinical diagnostics, however, utilize techniques involving surface plasmon resonance (SPR) and enzyme-linked immunosorbent assays (ELISAs), which are expensive and unfeasible in terms of portability. To develop a better, competitive biosensor, we must reduce the cost of one, or increase the sensitivity of the other. Electric fields are commonly utilized in microfluidic devices to manipulate particles, biomolecules, and cells. Applications in this area, however, are primarily limited to interfaces formed between immiscible interfaces. Miscible, liquid-liquid interfaces are common in microfluidic devices, and are easily reproduced with simple geometries. Here, we demonstrate the use of electrical fields at liquid-liquid electrical interfaces, known as fluidic dielectrophoresis, (fDEP) for biodetection in a microfluidic device. In this work, we apply an AC electric field across concurrent laminar streams with differing conductivities and permittivities to polarize the interface and induce a discernible, near-immediate, frequency-dependent interfacial tilt. We design this aqueous electrical interface, which becomes the biosensing “substrate,” to be intelligent – it “moves” only when a target of interest is present. This motion requires neither labels nor expensive electrical equipment, so the biosensor is inexpensive and portable, yet still capable of sensitive detection. Nanoparticles, due to their high surface-area-to-volume ratio, are often incorporated to enhance detection capabilities of schemes like SPR and fluorimetric assays. Most studies currently investigate binding at an immobilized solid-liquid or solid-gas interface, where particles are adsorbed onto a planar surface, functionalized with a receptor to create a reactive substrate, and subsequently flushed with a fluid or gas with the relevant analyte. These typically involve many preparation and rinsing steps, and are susceptible to surface fouling. Our microfluidic device is continuously flowing and renewing the “substrate,” and is thus not subject to fouling. In this work, we demonstrate the ability to electrokinetically detect biomolecules binding to functionalized nanoparticles at liquid-liquid interfaces using fDEP. In biotin-streptavidin experiments, we report binding detection limits on the order of 1-10 pM, without amplifying signals or concentrating samples. We also demonstrate the ability to detect this interfacial motion, and thus the presence of binding, using impedance spectroscopy, allowing this scheme to become non-optical, in addition to being label-free.

Keywords: biodetection, dielectrophoresis, microfluidics, nanoparticles

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Authors: Wonil Nam, Xiang Ren, Inyoung Kim, Masoud Agah, Wei Zhou

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Despite its ultrasensitive detection capability, surface-enhanced Raman spectroscopy (SERS) faces challenges as a quantitative biochemical analysis tool due to the significant dependence of local field intensity in hotspots on nanoscale geometric variations of plasmonic nanostructures. Therefore, despite enormous progress in plasmonic nanoengineering of high-performance SERS devices, it is still challenging to quantitatively correlate the measured SERS signals with the actual molecule concentrations at hotspots. A significant effort has been devoted to developing SERS calibration methods by introducing internal standards. It has been achieved by placing Raman tags at plasmonic hotspots. Raman tags undergo similar SERS enhancement at the same hotspots, and ratiometric SERS signals for analytes of interest can be generated with reduced dependence on geometrical variations. However, using Raman tags still faces challenges for real-world applications, including spatial competition between the analyte and tags in hotspots, spectral interference, laser-induced degradation/desorption due to plasmon-enhanced photochemical/photothermal effects. We show that electronic Raman scattering (ERS) signals from metallic nanostructures at hotspots can serve as the internal calibration standard to enable quantitative SERS analysis and improve biostatistical analysis. We perform SERS with Au-SiO₂ multilayered metal-insulator-metal nano laminated plasmonic nanostructures. Since the ERS signal is proportional to the volume density of electron-hole occupation in hotspots, the ERS signals exponentially increase when the wavenumber is approaching the zero value. By a long-pass filter, generally used in backscattered SERS configurations, to chop the ERS background continuum, we can observe an ERS pseudo-peak, IERS. Both ERS and SERS processes experience the |E|⁴ local enhancements during the excitation and inelastic scattering transitions. We calibrated IMRS of 10 μM Rhodamine 6G in solution by IERS. The results show that ERS calibration generates a new analytical value, ISERS/IERS, insensitive to variations from different hotspots and thus can quantitatively reflect the molecular concentration information. Given the calibration capability of ERS signals, we performed label-free SERS analysis of living biological systems using four different breast normal and cancer cell lines cultured on nano-laminated SERS devices. 2D Raman mapping over 100 μm × 100 μm, containing several cells, was conducted. The SERS spectra were subsequently analyzed by multivariate analysis using partial least square discriminant analysis. Remarkably, after ERS calibration, MCF-10A and MCF-7 cells are further separated while the two triple-negative breast cancer cells (MDA-MB-231 and HCC-1806) are more overlapped, in good agreement with the well-known cancer categorization regarding the degree of malignancy. To assess the strength of ERS calibration, we further carried out a drug efficacy study using MDA-MB-231 and different concentrations of anti-cancer drug paclitaxel (PTX). After ERS calibration, we can more clearly segregate the control/low-dosage groups (0 and 1.5 nM), the middle-dosage group (5 nM), and the group treated with half-maximal inhibitory concentration (IC50, 15 nM). Therefore, we envision that ERS calibrated SERS can find crucial opportunities in label-free molecular profiling of complicated biological systems.

Keywords: cancer cell drug efficacy, plasmonics, surface-enhanced Raman spectroscopy (SERS), SERS calibration

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