Layer-by-layer coating of bacteria with noble metal nanoparticles for surface-enhanced Raman scattering

A simple layer-by-layer method to coat the bacterial cells with gold and silver nanoparticles (AuNPs and AgNPs) for the acquisition of surface-enhanced Raman scattering (SERS) spectra is reported. First, the bacteria cell wall is coated with poly (allylamine hydrochloride) (PAH), a positively charged polymer, and then with citrate reduced Au or AgNPs. In order to increase the stability of the coating, another layer of PAH is prepared on the surface. The SEM and AFM images indicate that the nanoparticles are in the form of both isolated and aggregated nanoparticles on the bacterial wall. The coating of bacterial cells with AgNPs or AuNPs not only serves for their preparation for SERS measurement but also helps to visualize the coated of bacterial cells under the ordinary white-light microscope objective due to efficient light-scattering properties of Au and AgNPs. A comparative study single versus aggregates of bacterial cells is also demonstrated for possible single bacterial detection with SERS. The two bacteria that differ in shape and cell wall biochemical structure, Escherichia coli and Staphylococcus cohnii, Gram-negative and -positive, respectively, are used as models. The preliminary results reveal that the approach could be used for single bacterial cell identification. © 2009 Springer-Verlag

Functional artificial free-standing yeast biofilms

Here we report fabrication of artificial free-standing yeast biofilms built using sacrificial calcium carbonate-coated templates and layer-by-layer assembly of extracellular matrix-mimicking polyelectrolyte multilayers. The free-standing biofilms are freely floating multilayered films of oppositely charged polyelectrolytes and live cells incorporated in the polyelectrolyte layers. Such biofilms were initially formed on glass substrates of circular and ribbon-like shapes coated with thin layers of calcium carbonate microparticles. The templates were then coated with cationic and anionic polyelectrolytes to produce a supporting multilayered thin film. Then the yeast alone or mixed with various micro- and nanoparticle inclusions was deposited onto the multilayer composite films and further coated with outer polyelectrolyte multilayers. To detach the biofilms from the glass substrates the calcium carbonate layer was chemically dissolved yielding free-standing composite biofilms. These artificial biofilms to a certain degree mimic the primitive multicellular and colonial species. We have demonstrated the added functionality of the free-standing artificial biofilms containing magnetic, latex and silver micro- and nanoparticles. We have also developed "symbiotic" multicellular biofilms containing yeast and bacteria. This approach for fabrication of free-standing artificial biofilms can be potentially helpful in development of artificial colonial microorganisms composed of several different unicellular species and an important tool for growing cell cultures free of supporting substrates. © 2011 Elsevier B.V

Surface-Enhanced Raman Scattering to Evaluate Nanomaterial Cytotoxicity on Living Cells

© 2016 American Chemical Society.The increasing number of reports about false positive or negative results from conventional cytotoxicity assays of nanomaterials (NMs) suggests that more reliable NM toxicity assessment methods should be developed. Here, we report a novel approach for nanotoxicity evaluation based on surface-enhanced Raman spectroscopy (SERS). Three model NMs were tested on two model cell lines and the results were validated by WST-1 cytotoxicity assay and annexin V-FITC/propidium iodide (PI) staining as apoptosis-necrosis assay. The localization of nanoparticles (NPs) in the cells and the cellular conditions upon NP incubation were visualized by transmission electron microscopy (TEM) and enhanced dark-field (EDF) microscopy. SERS revealed a broader view on the consequences of cell-NM interactions compared to the conventional cytotoxicity assays where only one aspect of toxicity can be measured by one assay type. The results suggest that SERS can significantly contribute to the cytotoxicity evaluation bypassing NM or assay component-related complications with less effort

Binding and purification of plasmid DNA using multi-layered carbon nanotubes

We propose a new method for the separation of nucleic acids using multi-layered carbon nanotubes (CNTs) as an adsorbent. According to agarose gel electrophoresis, oxidized water-stable CNTs adsorb certain forms of nucleic acids, such as high molecular weight RNA, chromosomal DNA, linear and denatured forms of plasmid DNA. However, CNTs do not adsorb supercoiled form of plasmid DNA. Nucleic acids bound to CNTs can be readily removed by centrifugation whereas supercoiled plasmid DNA remains in solution. Upon the addition of divalent metal ions supercoiled plasmid DNA forms relatively stable complexes with CNTs due to chelation. Thus, new details about association of nucleic acids with CNTs were revealed and stoichiometry of the complexes was estimated. Our results can be used for fine purification of supercoiled plasmid DNA for gene therapy applications as well as manipulation of nucleic acids for biosensor design. © 2011 Elsevier B.V

Living fungi cells encapsulated in polyelectrolyte shells doped with metal nanoparticles

We report the layer-by-layer coating of living fungi cells (Saccharomyces cerevisiae and Trichoderma asperellum) with polyelectrolytes poly(allylamine hydrochloride)/sodium poly(styrene sulfonate) and bovine serum albumin/DNA and citrate-stabilized gold and silver nanoparticles. It was found that the nanoparticles were effectively incorporated between oppositely charged polyelectrolyte layers, modifying the topography and the roughness of cell walls. The formation of large aggregates of nanoparticles on the cell walls of encapsulated cells was shown. It was found that the encapsulated cells preserved their viability and the shells were soft enough to allow the growth of mycelium. The surface-enhanced Raman scattering (SERS) was used to investigate the biochemical environments of the gold and silver nanoparticles immobilized on the surface of T. asperellum conidia. The SERS spectra from encapsulated conidia and polyelectrolytes indicate that both gold and silver nanoparticles interact with cell walls from different locations, and nanoparticle - polyelectrolyte interaction is limited. The approach described in this paper might have potential applications in modification of living cells. © 2009 American Chemical Society

Lithium promotes long-term neurological recovery after spinal cord injury in mice by enhancing neuronal survival, gray and white matter remodeling, and long-distance axonal regeneration

Spinal cord injury (SCI) induces neurological deficits associated with long-term functional impairments. Since the current treatments remain ineffective, novel therapeutic options are needed. Besides its effect on bipolar mood disorder, lithium was reported to have neuroprotective activity in different neurodegenerative conditions, including SCI. In SCI, the effects of lithium on long-term neurological recovery and neuroplasticity have not been assessed. We herein investigated the effects of intraperitoneally administered lithium chloride (LiCl) on motor coordination recovery, electromyography (EMG) responses, histopathological injury and remodeling, and axonal plasticity in mice exposed to spinal cord transection. At a dose of 0.2, but not 2.0 mmol/kg, LiCl enhanced motor coordination and locomotor activity starting at 28 days post-injury (dpi), as assessed by a set of behavioral tests. Following electrical stimulation proximal to the hemitransection, LiCl at 0.2 mmol/kg decreased the latency and increased the amplitude of EMG responses in the denervated hindlimb at 56 dpi. Functional recovery was associated with reduced gray and white matter atrophy rostral and caudal to the hemitransection, increased neuronal survival and reduced astrogliosis in the dorsal and ventral horns caudal to the hemitransection, and increased regeneration of long-distance axons proximal and distal to the lesion site in mice receiving 0.2 mmol/kg, but not 2 mmol/kg LiCl, as assessed by histochemical and immunohistochemical studies combined with anterograde tract tracing. Our results indicate that LiCl induces long-term neurological recovery and neuroplasticity following SCI.TUBA ; Istanbul Medipol University ; Turkish Academy of Science

Nanostructured Silver Substrates With Stable and Universal SERS Properties: Application to Organic Molecules and Semiconductor Nanoparticles

Nanostructured silver films have been prepared by thermal deposition on silicon, and their properties as SERS substrates investigated. The optimal conditions of the post-growth annealing of the substrates were established. Atomic force microscopy study revealed that the silver films with relatively dense and homogeneous arrays of 60–80-nm high pyramidal nanoislands are the most efficient for SERS of both organic dye and inorganic nanoparticles analytes. The noticeable enhancement of the Raman signal from colloidal nanoparticles with the help of silver island films is reported for the first time

Towards a target label-free suboptimum oligonucleotide displacement-based detection system

A novel method for the future development of label-free DNA sensors is proposed here. The approach is based on the displacement of a labelled suboptimum mutated oligonucleotide hybridised with the immobilised biotin-capture probe. The target fully complementary to the biotin-capture probe can displace the labelled oligonucleotide causing a subsequent decrease of the signal that verifies the presence of the target. The decrease of signal was demonstrated to be proportional to the target concentration. A study of the hybridisation of mutated and complementary labelled oligonucleotides with an immobilised biotin-capture probe was carried out. Different kinetic and thermodynamic behaviour was observed for heterogeneous hybridisation of biotin-capture probe with complementary or suboptimum oligonucleotides. The displacement method evaluated colourimetrically achieved the objective of decreasing the response time from 1 h for direct hybridisation of 19-mer oligonucleotides in the direct enzyme-linked oligonucleotide assay (ELONA) to 5 min in the case of displacement detection in the micromolar concentration range