We report on our activities related to the development of surface enhanced Raman scattering (SERS) probes realized onto the optical fiber tip (OFT) through nanosphere lithography. In the first stage of our research, we adapted the nanosphere lithography to operate on the optical fiber tip, by assessing the process and demonstrating either the potentiality or the repeatability of the proposed nanopatterning approach. Successively, we investigated the ability of the manufactured structures on the fiber tip to act as SERS probes by measuring the SERS spectra in presence of a Biphenyl Thiol (BPT) monolayer. Firstly, we focused the attention on the samples shaped as closed packed array of nanospheres covered by gold. The analysis allowed us to identify the most promising SERS platform, exhibiting an Enhancement Factor (EF) of 4×105 and a SERS measurements variability lower than 10%. We addressed also the limitations related to the use of the same optical fiber for both illumination and light collection by selecting a commercial optical fiber exhibiting a suitable trade-off in terms of high excitation/collection efficiency and low silica background. Current activities are devoted to the investigation of other nanopatterns on the optical fiber tip (namely, Sparse Array of metallodielectric spheres) and the analysis of the probes response against different molecules.
In this work, we employed the Raman microscopy to study the internalization kinetics and spatial distribution of small interfering RNA (siRNA)-diatomite nanoparticles (DNPs) complex in human lung epidermoid carcinoma cell line (H1355) up to 72 h. Raman images are compared with confocal fluorescence microscopy results. The Raman analysis provides that the siRNA-DNPs are internalized and co-localized in lipid vesicles within 18 h, after that equilibrium is achieved.
Metasurfaces are two-dimensional structures, arrays of scatterers with subwavelength separation or optically thin planar films, allowing light manipulation and enabling specific changes of optical properties, as for example beam-steering, anomalous refraction and optical-wavefront shaping. Due to the fabrication simplicity, the metasurfaces offer an alternative to 3-D metamaterials and providing a novel method for optical elements miniaturization. It has been demonstrated that a metasurface can support Bound States in Continuum (BIC), that are resonant states by zero width, due to the interaction between trapped electromagnetic. Experimentally, this involves very narrow coupled resonances, with a high Q-factor and an extremely large field intensity enhancement, up to 6 orders of magnitude larger than the intensity of the incident beam. Here, we demonstrate that the field enhancement in proximity of the surface can be applied to boost fluorescence emission of probe molecules dispersed on the surface of a photonic crystal membrane fabricated in silicon nitride. Our results provide new solutions for light manipulation at the nanoscale, especially for sensing and nonlinear optics applications.
Semen analysis is widely used as diagnostic tool for assessing male fertility, controlling and managing the animal reproduction. The most important parameters measured in a semen analysis are the morphology and biochemical alterations. For obtaining such information, non-invasive, label-free and non-destructive techniques have to be used. Digital Holography (DH) combined with Raman Spectroscopy (RS) could represent the perfect candidate for a rapid, non-destructive and high-sensitive morphological and biochemical sperm cell analysis. In this study, DH-RS combined approach is used for a complete analysis of single bovine spermatozoa. High-resolution images of bovine sperm have been obtained by DH microscopy from the reconstruction of a single acquired hologram, highlighting in some cases morphological alterations. Quantitative 3D reconstructions of sperm head, both normal and anomalous, have been studied and an unexpected structure of the post-acrosomal region of the head has been detected. Such anomalies have been also confirmed by Raman imaging analysis, suggesting the protein vibrations as associated Raman marker of the defect.
Currently, a combination of technologies is typically required to identify and classify leukemia cells. These methods often lack the specificity and sensitivity necessary for early and accurate diagnosis. Here, we demonstrate the use of Raman spectroscopy to identify normal B cells, collected from healthy patients, and three ALL cell lines (RS4;11, REH and MN60 at different differentiation level, respectively). Raman markers associated with DNA and protein vibrational modes have been identified that exhibit excellent discriminating power for leukemia cell identification. Principal Component Analysis was finally used to confirm the significance of these markers for identify leukemia cells and classifying the data. The obtained results indicate a sorting accuracy of 96% between the three leukemia cell lines.
We developed a SERS biosensor based on gold fishnets fabricated by using e-beam lithography. This device is used for glycerophosphoinositol (GroPIns) molecule sensing. GroPIns is an abundant component of cell cytosol and high GroPIns levels have been reported in several tumour cells. We demonstrate that our SERS sensor is able to accurately and quantitatively determine the concentration of GroPIns. These results indicate that SERS may provide a novel platform technology to identify GroPIns profiles in disease pathogenesis.
The current study relates to a Raman spectroscopy-based method for addressing the problem of sex assessment in
mammals. A direct method for sex predetermination in animals is based on the X- and Y-bearing sperm cells sorting
before insemination. Our Raman spectroscope allows distinguishing and characterizing the difference between X- and
Y-bearing sperm cells by detecting and analyzing their Raman spectra in a non-invasive and non-destructive way.
Raman spectroscopy is a label-free and non-invasive method that measures the inelastic scattered light from a sample
giving insight into the vibration eigenmodes of the excited molecules. For these reasons, Raman spectroscopy has been
used as a powerful tool to investigate different biological tissues and living cells. In this paper, we present a Raman
spectroscopy-based method for sensitive biochemical characterization of bovine sperm cells. Importantly, by analysing
separate Raman spectra from the nucleus, acrosomale vesicle and tail of single sperm cells, we are able to identify
characteristic Raman features associated with DNA, protein and lipid molecular vibrations for discriminating among
different locations inside the cell with sub-micrometric resolution (∼0.3 μm). We demonstrate that our Raman
spectroscopy facilitates spectral assignment and increases detection sensitivity, opening the way for novel bio-imaging