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Optical tweezers exploit light–matter interactions to trap particles ranging from single atoms to micrometer-sized eukaryotic cells. For this reason, optical tweezers are a ubiquitous tool in physics, biology, and nanotechnology. Recently, the use of deep learning has started to enhance optical tweezers by improving their design, calibration, and real-time control as well as the tracking and analysis of the trapped objects, often outperforming classical methods thanks to the higher computational speed and versatility of deep learning. In this perspective, we show how cutting-edge deep learning approaches can remarkably improve optical tweezers, and explore the exciting, new future possibilities enabled by this dynamic synergy. Furthermore, we offer guidelines on integrating deep learning with optical trapping and optical manipulation in a reliable and trustworthy way.

Two-dimensional transition metal dichalcogenides, particularly MoS2, are interesting materials for many applications in aerospace research, radiation therapy and bioscience more in general. Since in many of these applications MoS2-based nanomaterials can be placed in an aqueous environment while exposed to ionizing radiation, both experimental and theoretical studies of their behaviour under these conditions is particularly interesting. Here, we study the effects of tiny imparted doses of 511 keV photons to MoS2 nanoflakes in water solution. To the best of our knowledge, this is the first study in which ionizing radiation on 2D-MoS2 occurs in water. Interestingly, we find that, in addition to the direct interaction between high-energy photons and nanoflakes, reactive chemical species, generated by γ-photons induced radiolysis of water, come into play a relevant role. A radiation transport Monte Carlo simulation allowed determining the elements driving the morphological and spectroscopical changes of 2D-MoS2, experimentally monitored by SEM microscopy, DLS, Raman and UV–vis spectroscopy, AFM, and X-ray photoelectron techniques. Our study demonstrates that radiolysis products affect the Molybdenum oxidation state, which is massively changed from the stable + 4 and + 6 states into the rarer and more unstable + 5. These findings will be relevant for radiation-based therapies and diagnostics in patients that are assuming drugs or contrast agents containing 2D-MoS2 and for aerospace biomedical applications of 2DMs investigating their actions into living organisms on space station or satellites. [ABSTRACT FROM AUTHOR]

Surface-enhanced Raman scattering (SERS) is of growing interest for a wide range of applications, especially for biomedical analysis, thanks to its sensitivity, specificity, and multiplexing capabilities. A crucial role for successful applications of SERS is played by the development of reproducible, efficient, and facile procedures for the fabrication of metal nanostructures (SERS substrates). Even more challenging is to extend the fabrication techniques of plasmonic nano-textures to atomic force microscope (AFM) probes to carry out tip-enhanced Raman spectroscopy (TERS) experiments, in which spatial resolution below the diffraction limit is added to the peculiarities of SERS. In this short review, we describe recent studies performed by our group during the last ten years in which novel nanofabrication techniques have been successfully applied to SERS and TERS experiments for studying bio-systems and molecular species of environmental interest. [ABSTRACT FROM AUTHOR]

The development of sensitive methods for the detection of endotoxin molecules, such as lipopolysaccharides (LPS), is essential for food safety and health control. Conventional analytical methods used for LPS detection are based on the pyrogen test, plating and culture-based methods, and the limulus amoebocyte lysate method (LAL). Alternatively, the development of reliable biosensors for LPS detection would be highly desirable to solve some critical issues, such as high cost and a long turnaround time. In this work, we present a label-free Surface-Enhanced Raman Spectroscopy (SERS)-based method for LPS detection in its free form. The proposed method combines the benefits of plasmonic enhancement with the selectivity provided by a specific anti-lipid A antibody (Ab). A high-enhancing nanostructured silver substrate was coated with Ab. The presence of LPS was quantitatively monitored by analyzing the changes in the Ab spectra obtained in the absence and presence of LPS. A limit of detection (LOD) and quantification (LOQ) of 12 ng/mL and 41 ng/mL were estimated, respectively. Importantly, the proposed technology could be easily expanded for the determination of other biological macromolecules. [ABSTRACT FROM AUTHOR]

The pursuit of environmentally friendly solvents has become an essential research topic in sustainable chemistry and nanomaterial science. With the need to substitute toxic solvents in nanofabrication processes becoming more pressing, the search for alternative solvents has taken on a crucial role in this field. Additionally, the use of toxic, non-economical organic solvents, such as N-methyl-2 pyrrolidone and dimethylformamide, is not suitable for all biomedical applications, even though these solvents are often considered as the best exfoliating agents for nanomaterial fabrication. In this context, the success of producing two-dimensional transition metal dichalcogenides (2D TMDs), such as MoS2 and WS2, with excellent captivating properties is due to the ease of synthesis based on environment-friendly, benign methods with fewer toxic chemicals involved. Herein, we report for the first time on the use of cyrene as an exfoliating agent to fabricate monolayer and few-layered 2D TMDs with a versatile, less time-consuming liquid-phase exfoliation technique. This bio-derived, aprotic, green and eco-friendly solvent produced a stable, surfactant-free, concentrated 2D TMD dispersion with very interesting features, as characterized by UV–visible and Raman spectroscopies. The surface charge and morphology of the fabricated nanoflakes were analyzed using ς-potential and scanning electron microscopy. The study demonstrates that cyrene is a promising green solvent for the exfoliation of 2D TMD nanosheets with potential advantages over traditional organic solvents. The ability to produce smaller-sized—especially in the case of WS2 as compared to MoS2—and mono/few-layered nanostructures with higher negative surface charge values makes cyrene a promising candidate for various biomedical and electronic applications. Overall, the study contributes to the development of sustainable and environmentally friendly methods for the production of 2D nanomaterials for various applications. [ABSTRACT FROM AUTHOR]

Nanoparticles (NPs) coated with hyaluronic acid (HA) seem to be increasingly promising for targeted therapy due to HA chemical versatility, which allows them to bind drugs of different natures, and their affinity with the transmembrane receptor CD-44, overexpressed in tumor cells. However, an essential aspect for clinical use of NPs is formulation stability over time. For these reasons, analytical techniques capable of characterizing their physico-chemical properties are needed. In this work, poly(lactide-co-glycolide) (PLGA) NPs with an average diameter of 100–150 nm, coated with a few 10 s of nm of HA, were synthesized. For stability characterization, two complementary investigative techniques were used: Dynamic Light Scattering (DLS) and Surface-Enhanced Raman Scattering (SERS) spectroscopy. The first technique provided information on size, polidispersity index, and zeta-potential, and the second provided a deeper insight on the NP surface chemicals, allowing distinguishing of HA-coated NPs from uncoated ones. Furthermore, in order to estimate formulation stability over time, NPs were measured and monitored for two weeks. SERS results showed a progressive decrease in the signal associated with HA, which, however, is not detectable by the DLS measurements. [ABSTRACT FROM AUTHOR]

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous pollutants that are typically released into the environment during the incomplete combustion of fossil fuels. Due to their relevant carcinogenicity, mutagenicity, and teratogenicity, it is urgent to develop sensitive and cost-effective strategies for monitoring them, especially in aqueous environments. Surface-enhanced Raman spectroscopy (SERS) can potentially be used as a reliable approach for this purpose, as it constitutes a valid alternative to traditional techniques, such as liquid and gas chromatography. Nevertheless, the development of an SERS-based platform for detection PAHs has so far been hindered by the poor adsorption of PAHs onto silver- and gold-based SERS-active substrates. To overcome this limitation, several research efforts have been directed towards the development of functionalized SERS substrates for the improvement of PAH adsorption. However, these strategies suffer from the interference that functionalizing molecules can produce in SERS detection. Herein, we demonstrate the feasibility of label-free detection of pyrene by using a highly porous 3D-SERS substrate produced by an inductively coupled plasma (ICP). Thanks to the coral-like nanopattern exhibited by our substrate, clear signals ascribable to pyrene molecules can be observed with a limit of detection of 23 nM. The observed performance can be attributed to the nanoporous character of our substrate, which combines a high density of hotspots and a certain capability of trapping molecules and favoring their adhesion to the Ag nanopattern. The obtained results demonstrate the potential of our substrates as a large-area, label-free SERS-based platform for chemical sensing and environmental control applications. [ABSTRACT FROM AUTHOR]

Suitable metal nanostructures may induce surface-enhanced Raman scattering (SERS) enhancement factors (EFs) large-enough to reach single-molecule sensitivity. However, the gap hot-spot EF probability density function (PDF) has the character of a long-tail distribution, which dramatically mines the reproducibility of SERS experiments. Herein, we carry out electrodynamic calculations based on a 3D finite element method of two plasmonic nanostructures, combined with Monte Carlo simulations of the EF statistics under different external conditions. We compare the PDF produced by a homodimer of nanoparticles with that provided by a self-similar trimer. We show that the PDF is sensitive to the spatial distribution of near-field enhancement specifically supported by the nanostructure geometry. Breaking the symmetry of the plasmonic system is responsible for inducing particular modulations of the PDF tail resembling a multiple Poisson distribution. We also study the influence that molecular diffusion towards the hottest hot-spot, or selective hot-spot targeting, might have on the EF PDF. Our results quantitatively assess the possibility of designing the response of a SERS substrate so as to contain the intrinsic EF PDF variance and significantly improving, in principle, the reproducibility of SERS experiments. [ABSTRACT FROM AUTHOR]

Although oocyte cryopreservation has great potentials in the field of reproductive technologies, it still is an open challenge in the majority of domestic animals and little is known on the biochemical transformation induced by this process in the different cellular compartments. Raman micro-spectroscopy allows the non-invasive evaluation of the molecular composition of cells, based on the inelastic scattering of laser photons by vibrating molecules. The aim of this work was to assess the biochemical modifications of both the zona pellucida and cytoplasm of vitrified/warmed in vitro matured bovine oocytes at different post-warming times. By taking advantage of Principal Component Analysis, we were able to shed light on the biochemical transformation induced by the cryogenic treatment, also pointing out the specific role of cryoprotective agents (CPs). Our results suggest that vitrification induces a transformation of the protein secondary structure from the α-helices to the β-sheet form, while lipids tend to assume a more packed configuration in the zona pellucida. Both modifications result in a mechanical hardening of this cellular compartment, which could account for the reduced fertility rates of vitrified oocytes. Furthermore, biochemical modifications were observed at the cytoplasmic level in the protein secondary structure, with α-helices loss, suggesting cold protein denaturation. In addition, a decrease of lipid unsaturation was found in vitrified oocytes, suggesting oxidative damages. Interestingly, most modifications were not observed in oocytes exposed to CPs, suggesting that they do not severely affect the biochemical architecture of the oocyte. Nevertheless, in oocytes exposed to CPs decreased developmental competence and increased reactive oxygen species production were observed compared to the control. A more severe reduction of cleavage and blastocyst rates after in vitro fertilization was obtained from vitrified oocytes. Our experimental outcomes also suggest a certain degree of reversibility of the induced transformations, which renders vitrified oocytes more similar to untreated cells after 2 h warming. [ABSTRACT FROM AUTHOR]

We numerically investigate the electromagnetic field localization in a two-dimensional photonic quasicrystal generated with a holographic tiling. We demonstrate that light confinement can be induced into an air mirrorless cavity by the inherent symmetry of the spatial distribution of the dielectric scatterers forming the side walls of the open cavity. Furthermore, the propagation direction can be controlled by suitable designs of the structure. This opens up new avenues for designing photonic materials and devices. [ABSTRACT FROM AUTHOR]

Conjunctival microvilli are microscopic cellular membrane protrusions on apical epithelial cells, which increase the surface area available for tear adherence. Pathological alterations of microvilli structure affect the tear film stability and, conversely, dysfunctions of tear film composition can lead to a suffering epithelium (dry-eye syndrome). In this work we propose the use of micro-Raman analysis to reveal conjunctival microvilli abnormalities. Samples were obtained by impression cytology from patients by different stage of dry-eye syndrome. Our experimental outcomes demonstrate that Raman analysis, combined with the use of Principal Component Analysis, is able to detect different stages of microvilli reduction. Globally, these results hold promise for the use of Raman analysis for an objective, effective, non-invasive and potentially also in-vivo analysis of the conjunctiva in all the cases of microvilli-related ocular pathologies. [ABSTRACT FROM AUTHOR]

Bacterial spores are surrounded by the coat, a multilayered shell that contributes in protecting the genome during stress conditions. In Bacillus subtilis, the model organism for spore formers, the coat is composed by about seventy different proteins, organized into four layers by the action of several regulatory proteins. A major component of this regulatory network, CotE, is needed to assemble the outer coat and develop spores fully resistant to lysozyme and able to germinate efficiently. Another regulator, CotH, is controlled by CotE and is present in low amounts both during sporulation and in mature spores. In spite of this CotH controls the assembly of at least nine outer coat proteins and cooperates with CotE in producing fully resistant and efficiently germinating spores. In order to improve our understanding of CotH role in spore formation, we over-produced CotH by placing its coding region under the control of a promoter stronger than its own promoter but with a similar timing of activity during sporulation. Over-production of CotH in an otherwise wild type strain did not cause any major effect, whereas in a cotE null background a partial recovery of the phenotypes associated to the cotE null mutation was observed. Western blot, fluorescence microscopy and Surface-Enhanced Raman Scattering spectroscopy data indicate that, in the absence of CotE, over-production of CotH allowed the formation of spores overall resembling wild type spores and carrying in their coat some CotE−/CotH-dependant proteins. Our results suggest that the B. subtilis spore differentiation programme is flexible, and that an increase in the amount of a regulatory protein can replace a missing partner and partially substitute its function in the assembly of the spore coat. [ABSTRACT FROM AUTHOR]

Acanthamoeba keratitis is a rare but serious corneal disease, often observed in contact lens wearers. Clinical treatment of infected patients frequently involves the use of polyhexamethylene biguanide (PHMB), a polymer used as a disinfectant and antiseptic, which is toxic also for the epithelial cells of the cornea. Prompt and effective diagnostic tools are hence highly desiderable for both starting early therapy and timely suspension of the treatment. In this work we use Raman microspectroscopy to analyse in vitro a single Acanthamoeba cell in cystic phase. In particular, we investigate the effect of PHMB at the single-cell level, providing useful information on both the underlying biochemical mechanism and the time frame for Acanthamoeba eradication in ocular infections. Furthermore, we demonstrate that Raman spectroscopy, in conjunction with standard multivariate analysis methods, allows discriminating between live and dead Acanthamoebas, which is fundamental to optimizing patients’ treatment. [ABSTRACT FROM AUTHOR]

We studied the mechanical properties of living starfish oocytes belonging to two species, Astropecten Auranciacus and Asterina pectinifera, over a wide range of timescales. We monitored the Brownian motion of microspheres injected in the cytoplasm using laser particle-tracking (LPT) and video multiple-particle-tracking (MPT) techniques, to explore high- and low-frequency response ranges, respectively. The analysis of the mean-square-displacements (MSD) allowed us to characterize the samples on different timescales. The MSD behavior is explained by three power-law exponents: for short times ( τ < 1 ms) it reflects the semiflexible behavior of the actin network; for intermediate timescales (1 ms < τ < 1 s) it is similar to that of a soft-glass material; finally for long times ( τ > 1 s) it behaves mainly like a viscous medium. We computed and compared the viscoelastic moduli using a recently proposed model describing the frequency response of the cell material. The large fluctuations found in the MSD over hundreds of trajectories indicate and confirm the significant cytoplasm heterogeneity. (© 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [ABSTRACT FROM AUTHOR]

This review presents the development of a Raman Tweezers system for detecting hemoglobin-related blood disorders at a single cell level. The study demonstrates that the molecular fingerprint insight provided by Raman analysis holds great promise for distinguishing between healthy and diseased cells in the field of biomedicine. Herein a Raman Tweezers system has been applied to investigate the effects of thalassemia, a blood disease quite diffuse in the Mediterranean Sea region. By resonant excitation of hemoglobin Raman bands, we examined the oxygenation capability of normal, alpha- and beta-thalassemic erythrocytes. A reduction of this fundamental red blood cell function, particularly severe for beta-thalassemia, has been found. Raman spectroscopy was also used to draw hemoglobin distribution inside single erythrocytes; the results confirmed the characteristic anomaly (target shape), occurring in thalassemia and some other blood disorders. The success of resonance Raman spectroscopy for thalassemia detection reported in this review provide an interesting starting point to explore the application of a Raman Tweezers system in the analysis of several blood disorders. [ABSTRACT FROM AUTHOR]