E-Protein Protonation Titration-Induced Single-Particle Chemical Force Spectroscopy for Microscopic Understanding and pI Estimation of Infectious DENV.

The ionization state of amino acids on the outer surface of a virus regulates its physicochemical properties toward the sorbent surface. Serologically different strains of the dengue virus (DENV) show different extents of infectivity depending upon their interactions with a receptor on the host cell. To understand the structural dependence of E-protein protonation over its sequence dependence, we have followed E-protein titration kinetics both experimentally and theoretically for two differentially infected dengue serotypes, namely, DENV-2 and DENV-4. We have performed E-protein protonation titration-induced single-particle chemical force spectroscopy using an atomic force microscope (AFM) to measure the surface chemistry of DENV in physiological aqueous solutions not only to understand the charge distribution dynamics on the virus surface but also to estimate the isoelectric point (pI) accurately for infectious dengue viruses. Cryo-EM structure-based theoretical pI calculations of the DENV-2 surface protein were shown to be consistent with the evaluated pI value from force spectroscopy measurements. We also highlighted here the role of the microenvironment around the titrable residues (in the 3D-folded structure of the protein) in altering the pKa. This is a comprehensive study to understand how the cumulative charge distribution on the outer surface of a specific serotype of DENV regulates a prominent role of infectivity over minute changes at the genetic level.

UBR7 in concert with EZH2 inhibits the TGF-ß signaling leading to extracellular matrix remodeling.

The intricate interplay between resident cells and the extracellular matrix (ECM) profoundly influences cancer progression. In triple-negative breast cancer (TNBC), ECM architecture evolves due to the enrichment of lysyl oxidase, fibronectin, and collagen, promoting distant metastasis. Here we uncover a pivotal transcription regulatory mechanism involving the epigenetic regulator UBR7 and histone methyltransferase EZH2 in regulating transforming growth factor (TGF)-ß/Smad signaling, affecting the expression of ECM genes. UBR7 loss leads to a dramatic reduction in facultative heterochromatin mark H3K27me3, activating ECM genes. UBR7 plays a crucial role in matrix deposition in adherent cancer cells and spheroids, altering collagen content and lysyl oxidase activity, directly affecting matrix stiffness and invasiveness. These findings are further validated in vivo in mice models and TNBC patients, where reduced UBR7 levels are accompanied by increased ECM component expression and activity, leading to fibrosis-mediated matrix stiffness. Thus, UBR7 is a master regulator of matrix stiffening, influencing the metastatic potential of TNBC.

Popcorn-shaped magnetic core-plasmonic shell multifunctional nanoparticles for the targeted magnetic separation and enrichment, label-free SERS imaging, and photothermal destruction of multidrug-resistant bacteria.

Over the last few years, one of the most important and complex problems facing our society is treating infectious diseases caused by multidrug-resistant bacteria (MDRB), by using current market-existing antibiotics. Driven by this need, we report for the first time the development of the multifunctional popcorn-shaped iron magnetic core-gold plasmonic shell nanotechnology-driven approach for targeted magnetic separation and enrichment, label-free surface-enhanced Raman spectroscopy (SERS) detection, and the selective photothermal destruction of MDR Salmonella DT104. Due to the presence of the "lightning-rod effect", the core-shell popcorn-shaped gold-nanoparticle tips provided a huge field of SERS enhancement. The experimental data show that the M3038 antibody-conjugated nanoparticles can be used for targeted separation and SERS imaging of MDR Salmonella DT104. A targeted photothermal-lysis experiment, by using 670 nm light at 1.5 W cm(-2) for 10 min, results in selective and irreparable cellular-damage to MDR Salmonella. We discuss the possible mechanism and operating principle for the targeted separation, label-free SERS imaging, and photothermal destruction of MDRB by using the popcorn-shaped magnetic/plasmonic nanotechnology.

Theranostic magnetic core-plasmonic shell star shape nanoparticle for the isolation of targeted rare tumor cells from whole blood, fluorescence imaging, and photothermal destruction of cancer.

Cancer is one of the most life-threatening diseases, which causes 7.6 million deaths and around 1 trillion dollars economic loss every year. Theranostic materials are expected to improve early detection and safe treatment through personalized medicine. Driven by the needs, we report the development of a theranostic plasmonic shell-magnetic core star shape nanomaterial based approach for the targeted isolation of rare tumor cells from the whole blood sample, followed by diagnosis and photothermal destruction. Experimental data with whole blood sample spiked with SK-BR-3 cancer cell shows that Cy3 attached S6 aptamer conjugated theranostic plasmonic/magnetic nanoparticles can be used for fluorescence imaging and magnetic separation even in 0.001% mixtures. A targeted photothermal experiment using 1064 nm near-IR light at 2-3 W/cm(2) for 10 min resulted in selective irreparable cellular damage to most of the SK-BR-3 cancer cells. We discuss the possible mechanism and operating principle for the targeted imaging, separation, and photothermal destruction using theranostic magnetic/plasmonic nanotechnology. After the optimization of different parameters, this theranostic nanotechnology-driven assay could have enormous potential for applications as contrast agent and therapeutic actuators for cancer.

A surface enhanced Raman scattering probe for highly selective and ultra sensitive detection of iodide in water and salt samples.

Iodine is a biophilic and essential trace element for all life and especially for vertebrates, which require it to produce indispensable thyroid hormones in their thyroid glands. As a result, the adequate measurement of iodine in water and food samples is crucial to lead a healthy life. Motivated by its importance, this is the first time in the literature that the highly selective and ultra sensitive (30 ppt limit) surface enhanced Raman scattering (SERS)-based detection of iodide ions (I(-)) from environmental and food samples has been reported. The desired sensitivity and selectivity has been achieved by measuring the change in the SERS intensity originating from Rh6G-adsorbed 30 nm gold nanoparticles (GNPs) upon the addition of I(-). The strong chemical affinity offered by I(-) towards the gold surface results in extra negative charge being deposited on it. As a result, the GNP surface attracts a greater number of positively charged Rh6G molecules and induces a marked increase in the number of hot spots through aggregation, providing a significant enhancement of the Raman signal intensity. The oxidation of I(-) to molecular iodine (I(2)) by hydrogen peroxide (H(2)O(2)) is employed for the successful screening of the bromide ion (Br(-)) which shows substantial interference at higher concentrations.

Nanomaterials for targeted detection and photothermal killing of bacteria.

Despite the modern treatment processes, contamination of food, water and medical equipment by pathogenic bacteria is very common in this world. Since the last two decades, one of the most important and complex problems our society has been facing is that several human pathogens became resistant to most of the clinically approved antibiotics. Recent advancement in nanoscience and nanotechnology has expanded our ability to design and construct nanomaterials with targeting, therapeutic, and diagnostic functions. These multifunctional materials have attracted our attention to be used as the promising tool for selective bacteria sensing and therapy without the current drugs. This tutorial review provides the basic concepts and critical properties of the different nanostructures that are useful for the pathogen detection and photothermal applications. In addition, bio-conjugated nanomaterial based strategies have been discussed with the aim to provide readers an overview of exciting opportunities and challenges in this field.

Resultant inward imbalanced seeding force (RIISF)-induced concave gold nanostar (CAuNS) for non-enzymatic electrocatalytic detection of serotonin and Kynurenine in human serum.

CTAC-based gold nanoseed-induced concave curvature evolution of surface boundary planes from concave gold nanocube (CAuNC) to concave gold nanostar (CAuNS) has been achieved by a novel synthetic methodology simply by controlling the extent of seed used and hence the generated 'Resultant Inward Imbalanced Seeding Force (RIISF)'. The resultant CAuNS shows an excellent enhancement in catalytic activity compared to CAuNC and other intermediates as a function of curvature-induced anisotropy. Detailed characterization evaluates the presence of an enhanced number of multiple defect sites, high energy facets, larger surface area, and roughened surface which ultimately results in an increased mechanical strain, coordinately unsaturation, and multifacet-oriented anisotropic behavior suitable for positive influence on the binding affinity of CAuNSs. While different crystalline and structural parameters improve their catalytic activity, the resultant uniform three-dimensional (3D) platform shows comparatively easy pliability and well absorptivity on the glassy carbon electrode surface for increased shelf life, a uniform structure to confine a large extent of stoichiometric systems, and long-term stability under ambient conditions for making this newly developed material a unique nonenzymatic scalable universal electrocatalytic platform. With the help of various electrochemical measurements, the ability of the platform has been established by performing highly specific and sensitive detection of the two most important human bio messengers: Serotonin (STN) and Kynurenine (KYN) which are metabolites of L-Tryptophan in the human body system. The present study mechanistically surveys the role of seed-induced RIISF-modulated anisotropy in controlling the catalytic activity which offers a universal 3D electrocatalytic sensing tenet by an electrocatalytic approach.

Development of a long-range surface-enhanced Raman spectroscopy ruler.

Optical-ruler-based distance measurements are essential for tracking biomolecular processes in a wide range of analytical biochemical applications. The normally used Förster resonance energy transfer (FRET) ruler is not useful for investigating distance-dependent properties when distances are more than 10 nm. Driven by this limitation, we have developed a long-range surface-enhanced Raman spectroscopy (SERS) optical ruler using oval-shaped gold nanoparticles and Rh6G dye-modified rigid, variable-length double-strand DNAs. The bifunctional rigid dsDNA molecule serves as the SERS-active ruler. Our experimental results show that one can tune the length of the SERS ruler between 8 and ∼18 nm by choosing the size of the oval-shaped gold nanoparticles. A possible mechanism for our observed distance-dependent SERS phenomenon is discussed using the Gersten and Nitzan model. Ultimately, our long-range SERS molecular rulers can be an important step toward understanding distance-dependent biological processes.

Highly efficient SERS substrate for direct detection of explosive TNT using popcorn-shaped gold nanoparticle-functionalized SWCNT hybrid.

This paper reports for the first time the development of a large-scale SERS substrate from a popcorn-shaped gold nanoparticle-functionalized single walled carbon nanotubes hybrid thin film for the selective and highly sensitive detection of explosive TNT material at a 100 femtomolar (fM) level.

A Mechanoelastic Glimpse on Hyaluronan-Coated Extracellular Vesicles.

Cancer cells secrete extracellular vesicles (EVs) covered with a carbohydrate polymer, hyaluronan (HA), linked to tumor malignancy. Herein, we have unravelled the contour lengths of HA on a single cancer cell-derived EV surface using single-molecule force spectroscopy (SMFS), which divulges the presence of low molecular weight HA (LMW-HA < 200 kDa). We also discovered that these LMW-HA-EVs are significantly more elastic than the normal cell-derived EVs. This intrinsic elasticity of cancer EVs could be directly allied to the LMW-HA abundance and associated labile water network on EV surface as revealed by correlative SMFS, hydration dynamics with fluorescence spectroscopy, and molecular dynamics simulations. This method emerges as a molecular biosensor of the cancer microenvironment.

Long-range nanoparticle surface-energy-transfer ruler for monitoring photothermal therapy response.

A recent gold nanotechnology-driven approach opens up a new possibility for the destruction of cancer cells through photothermal therapy. Ultimately, photothermal therapy may enter into clinical therapy and, as a result, there is an urgent need for techniques to monitor the tumor response to therapy. Driven by this need, a nanoparticle surface-energy-transfer (NSET) approach to monitor the photothermal therapy process by measuring a simple fluorescence intensity change is reported. The fluorescence intensity change is due to the light-controlled photothermal release of single-stranded DNA/RNA via dehybridization during the therapy process. Time-dependent results show that just by measuring the fluorescence intensity change, the photothermal therapy response during the therapy process can be monitored. The possible mechanism and operating principle of the NSET assay are discussed. Ultimately, this NSET assay could have enormous potential applications in rapid, on-site monitoring of the photothermal therapy process, which is critical to providing effective treatment of cancer and multidrug-resistant bacterial infections.

A label-free gold-nanoparticle-based SERS assay for direct cyanide detection at the parts-per-trillion level.

Cyanide is an extremely toxic lethal poison known to humankind. Developing rapid, highly sensitive, and selective detection of cyanide from water samples is extremely essential for human life safety. Driven by the need, here we report a gold-nanoparticle-based label-free surface-enhanced Raman spectroscopy (SERS) system for highly toxic cyanide ion recognition in parts-per-trillion level and to examine gold-nanoparticle-cyanide interaction. We have shown that the SERS assay can be used to probe the gold nanoparticle dissociation process in the presence of cyanide ions. Our experimental data indicates that gold-nanoparticle-based SERS can detect cyanide from a water sample at the 110 ppt level with excellent discrimination against other common anions and cations. The results also show that the SERS probe can be used to detect cyanide from environmental samples.

Impact of porous nanomaterials on inhibiting protein aggregation behaviour.

Aggregation of intrinsically disordered as well as the ordered proteins under certain premises or physiological conditions leads to pathological disorder. Here we have presented a detailed investigation on the effect of a porous metallic (Au) and a non-metallic (Si) nanomaterial on the formation of ordered (fiber-like/amyloid) and disordered (amorphous) aggregates of proteins. Porous nanogold (PNG) was found to reduce the amyloid aggregation of insulin but does not have much impact on the lag phase in the aggregation kinetics, whereas porous nano-silica (PNS) was found both to decrease the amount of aggregation as well as prolong the lag phase of amyloid fiber formation from insulin. On the other hand, both the porous nanoparticles are found to decrease the extent of amorphous aggregation (with slight improvement for PNS) of pathogenic huntingtin (Htt) protein in Huntington's disease cell model. This is a noted direct observation in controlling and understanding protein aggregation diseases which may help us to formulate nanotherapeutic drugs for future clinical applications.

Gold nano-popcorn-based targeted diagnosis, nanotherapy treatment, and in situ monitoring of photothermal therapy response of prostate cancer cells using surface-enhanced Raman spectroscopy.

Prostate cancer is the second leading cause of cancer-related death among the American male population, and the cost of treating prostate cancer patients is about $10 billion/year in the United States. Current treatments are mostly ineffective against advanced-stage prostate cancer and are often associated with severe side effects. Driven by these factors, we report a multifunctional, nanotechnology-driven, gold nano-popcorn-based surface-enhanced Raman scattering (SERS) assay for targeted sensing, nanotherapy treatment, and in situ monitoring of photothermal nanotherapy response during the therapy process. Our experimental data show that, in the presence of LNCaP human prostate cancer cells, multifunctional popcorn-shaped gold nanoparticles form several hot spots and provide a significant enhancement of the Raman signal intensity by several orders of magnitude (2.5 × 10(9)). As a result, it can recognize human prostate cancer cells at the 50-cells level. Our results indicate that the localized heating that occurs during near-infrared irradiation can cause irreparable cellular damage to the prostate cancer cells. Our in situ time-dependent results demonstrate for the first time that, by monitoring SERS intensity changes, one can monitor photothermal nanotherapy response during the therapy process. Possible mechanisms and operating principles of our SERS assay are discussed. Ultimately, this nanotechnology-driven assay could have enormous potential applications in rapid, on-site targeted sensing, nanotherapy treatment, and monitoring of the nanotherapy process, which are critical to providing effective treatment of cancer.

Rapid colorimetric identification and targeted photothermal lysis of Salmonella bacteria by using bioconjugated oval-shaped gold nanoparticles.

Salmonella bacteria are the major cause for the infection of 16 million people worldwide with typhoid fever each year. Antibiotic-resistant Salmonella strains have been isolated from various food products. As a result, the development of ultrasensitive sensing technology for detection and new approaches for the treatment of infectious bacterial pathogens that do not rely on traditional therapeutic regimes is very urgent for public health, food safety, and the world economy. Driven by this need, we report herein a nanotechnology-driven approach that uses antibody-conjugated oval-shaped gold nanoparticles to selectively target and destroy pathogenic bacteria. Our experiments have shown the use of a simple colorimetric assay, with an anti-salmonella antibody conjugated to oval-shaped gold nanoparticles, for the label-free detection of S. typhimurium with an excellent detection limit (10(4) bacteria per mL) and high selectivity over other pathogens. When bacteria conjugated to oval-shaped gold nanoparticles were exposed to near-infrared radiation, a highly significant reduction in bacterial cell viability was observed due to photothermal lysis. Ideally, this nanotechnology-based assay would have enormous potential for rapid, on-site pathogen detection to avoid the distribution of contaminated foods.

Effect of Surface Coating on the Toxicity of Silver Nanomaterials on Human Skin Keratinocytes.

As nanotechnology field continues to develop, assessing nanoparticle toxicity is very important for advancing nanoparticles for daily life application. In this Letter, we report the effect of surface coating on cyto, geno and photo-toxicity of silver nanomaterials of different shapes on human skin HaCaT keratinocytes. We found that the citrate coated colloidal silver nanoparticles at 100 µg/mL level are not geno-, cyto- and phtotoxic. On the other hand, citrate coated powder form of the silver nanoparticles are toxic. We have demonstrated that coating of the silver nanoparticles with a biodegradable polymer prevents the toxicity of the powder. Toxicity mechanism has been discussed.

Bimetallic gold-silver nanoparticles mediate bacterial killing by disrupting the actin cytoskeleton MreB.

The actin cytoskeleton is required for the maintenance of the cell shape and viability of bacteria. It remains unknown to which extent nanoparticles (NPs) can orchestrate the mechanical instability by disrupting the cytoskeletal network in bacterial cells. Our work demonstrates that Au-Ag NPs disrupt the bacterial actin cytoskeleton specifically, fluidize the inner membrane and lead to killing of bacterial cells. In this study, we have tried to emphasize on the key parameters important for NP-cell interactions and found that the shape, specific elemental surface localization and enhanced electrostatic interaction developed due to the acquired partial positive charge by silver atoms in the aggregated NPs are some of the major factors contributing towards better NP interactions and subsequent cell death. In vivo studies in bacterial cells showed that the NPs exerted a mild perturbation of the membrane potential. However, its most striking effect was on the actin cytoskeleton MreB resulting in morphological changes in the bacterial cell shape from rods to predominantly spheres. Exposure to NPs resulted in the delocalization of MreB patches from the membrane but not the tubulin homologue FtsZ. Concomitant with the redistribution of MreB localization, a dramatic increase of membrane fluid regions was observed. Our studies reveal for the first time that Au-Ag NPs can mediate bacterial killing and disrupt the actin cytoskeletal functions in bacteria.

Identification of Biomarker Hyaluronan on Colon Cancer Extracellular Vesicles Using Correlative AFM and Spectroscopy.

Extracellular vesicles (EVs), naturally occurring nanosized vesicles secreted from cells, are essential for intercellular communication. They carry unique biomolecules on the surface or interior that are of great interest as biomarkers for various pathological conditions such as cancer. In this work, we use high-resolution atomic force microscopy (AFM) and spectroscopy (AFS) techniques to demonstrate differences between EVs derived from colon cancer cells and colon epithelial cells at the single-vesicle level. We observe that EV populations are significantly increased in the cancer cell media compared to the normal cell EVs. We show that both EVs display an EV marker, CD9, while EVs derived from the cancer cells are slightly higher in density. Hyaluronan (HA) is a nonsulfated glycosaminoglycan linked to malignant tumor growth according to recent reports. Interestingly, at the single-vesicle level, colon cancer EVs exhibit significantly increased HA surface densities compared to the normal EVs. Spectroscopic measurements such as Fourier transform infrared (FT-IR), circular dichroism (CD), and Raman spectroscopy unequivocally support the AFM and AFS measurements. To our knowledge, it represents the first report of detecting HA-coated EVs as a potential colon cancer biomarker. Taken together, this sensitive approach will be useful in identifying biomarkers in the early stages of detection and evaluation of cancer.

Differential flexibility leading to crucial microelastic properties of asymmetric lipid vesicles for cellular transfection: A combined spectroscopic and atomic force microscopy studies.

The role of microscopic elasticity of nano-carriers in cellular uptake is an important aspect in biomedical research. Herein we have used AFM nano-indentation force spectroscopy and Förster resonance energy transfer (FRET) measurements to probe microelastic properties of three novel cationic liposomes based on di-alkyl dihydroxy ethyl ammonium chloride based lipids having asymmetry in their hydrophobic chains (Lip1818, Lip1814 and Lip1810). AFM data reveals that symmetry in hydrophobic chains of a cationic lipid (Lip1818) imparts higher rigidity to the resulting liposomes than those based on asymmetric lipids (Lip1814 and Lip1810). The stiffness of the cationic liposomes is found to decrease with increasing asymmetry in the hydrophobic lipid chains in the order of Lip1818 > Lip1814 > lip1810. FRET measurements between Coumarin 500 (Donor) and Merocyanine 540 (Acceptor) have revealed that full width at half-maxima (hw) of the probability distribution (P(r)) of donor-acceptor distance (r), increases in an order Lip1818 < Lip1814 < Lip1810 with increasing asymmetry of the hydrophobic lipid chains. This increase in width (hw) of the donor-acceptor distance distributions is reflective of increasing flexibility of the liposomes with increasing asymmetry of their constituent lipids. Thus, the results from AFM and FRET studies are complementary to each other and indicates that an increase in asymmetry of the hydrophobic lipid chains increases elasticity and or flexibility of the corresponding liposomes. Cell biology experiments confirm that liposomal flexibility or rigidity directly influences their cellular transfection efficiency, where Lip1814 is found to be superior than the other two liposomes manifesting that a critical balance between flexibility and rigidity of the cationic liposomes is key to efficient cellular uptake. Taken together, our studies reveal how asymmetry in the molecular architecture of the hydrophobic lipid chains influences the microelastic properties of the liposomes, and hence, their cellular uptake efficiency.

Gold nanoparticle based label-free SERS probe for ultrasensitive and selective detection of trinitrotoluene.

TNT is one of the most commonly used nitro aromatic explosives used for landmine and military purpose. Due to the significant detrimental effects, contamination of soil and groundwater with TNT is the major concern. Driven by the need to detect trace amounts of TNT from environmental samples, this article demonstrates for the first time a highly selective and ultra sensitive, cysteine modified gold nanoparticle based label-free surface enhanced Raman spectroscopy (SERS) probe, for TNT recognition in 2 pico molar (pM) level in aqueous solution. Due to the formation of Meisenheimer complex between TNT and cysteine, gold nanoparticles undergo aggregation in the presence of TNT via electrostatic interaction between Meisenheimer complex bound gold nanoparticle and cysteine modified gold nanoparticle. As a result, it formed several hot spots and provided a significant enhancement of the Raman signal intensity by 9 orders of magnitude through electromagnetic field enhancements. A detailed mechanism for termendous SERS intensity change has been discussed. Our experimental results show that TNT can be detected quickly and accurately without any dye tagging in lower pM level with excellent discrimination against other nitro compounds and heavy metals.