Intracavity optical tweezers have been proven successful for trapping microscopic particles at very low average power intensity – much lower than the one in standard optical tweezers. This feature makes them particularly promising for the study of biological samples. The modeling of such systems, though, requires time-consuming numerical simulations that affect its usability and predictive power. With the help of machine learning, we can overcome the numerical bottleneck – the calculation of optical forces, torques, and losses – reproduce the results in the literature and generalize to the case of counterpropagating-beams intracavity optical trapping.
Intracavity optical tweezers are a powerful tool to trap microparticles in water using the nonlinear feedback effect produced by the particle motion when it is trapped inside the laser cavity. In such systems two configurations are possible: a single-beam configuration and counterpropagating one. A removable isolator allows to switch between these configurations by suppressing one of the beams. Trapping a particle in the counterpropagating configuration, the measure of the optical power shows a feedback effect for each beam, that is present also when the two beams are misaligned and the trapped particle periodically jumps between them.
Detection of analytes in aqueous solution with high specificity and sensitivity is of paramount importance in many fields of science, ranging from biomedicine, environmental control, and food quality assessment. Surface-enhanced Raman scattering (SERS) has proven to be a cutting-edge analytical technique for this purpose, by combining the high selectivity of Raman features with the high sensitivity deriving from the plasmonic amplification of Raman signals. Herein, we report a facile and quite effective approach to fabricate large-area Ag-based SERS substrates, exhibiting a porous, coral-like nanotexture. Due to their intrinsic large surface-area and high hot-spot density, the produced substrates appear quite promising for the detection of analytes at trace levels. The nanoporous substrates are produced by Solid-State Dewetting (SSD) of thin Ag-films. In particular, ~30 nm thickness Ag-films are first deposited on glass coverslips by magnetron sputtering. Then, marked roughening is induced by exposing the films to an Inductively Coupled Plasma (ICP) discharge, using synthetic air as feeding gas. The performances of our SERS substrates are characterized in terms of morphology and enhancement factor using CV as probe molecule.
The phenomenon of inclusions or microvacuoles in intraocular lenses (IOL), often referred to glistenings due to their appearance when visualized in slit-lamp exams, is main cause of decreased visual in people after IOL implantation. For this reason, there is a huge request by the market of new polymers able to reduce, or even eliminate, the formation of such microvacuoles. In such frame, the use of advanced optical techniques, able to provide a deeper insight on the glistering formation, is strongly required. In particular, Raman spectroscopy (RS) is ideally suited for the analysis of polymers, due to its well-know sensitivity to highly polarizable chemical groups, commonly found in the polymer chains backbones. Moreover, the combination of RS with optical microscopy (Raman micro-spectroscopy) paves the way for real, information-rich chemical mapping of polymeric materials (Raman imaging). In this paper, we analyze the formation of microvacuoles in IOLs following a thermal treatment. In particular, we performed a chemical mapping of a single microvacuole, which allowed us to infer on its effective chemical composition. In order to investigate on the reversibility of glistenings formation, this analysis was repeated as function of time after thermal treatment, in different IOL environments. It turns out that this phenomenon is partially reversible, with an almost complete disappearance of microvacuoles in a dry environment.
Understanding of the complex interactions of molecules at biological interfaces is a fundamental issue in biochemistry, biotechnology as well as biomedicine. A plethora of biological processes are ruled by the molecular texture of cellular membrane: cellular communications, drug transportations and cellular recognition are just a few examples of such chemically-mediated processes. Tip-Enhanced Raman Scattering (TERS) is a novel, Raman-based technique which is ideally suited for this purpose. TERS relies on the combination of scanning probe microscopy and Raman spectroscopy. The basic idea is the use of a metalled tip as a sort of optical nano-antenna, which gives place to SERS effect close to the tip end. Herein, we present the application of TERS to analyze the surface of Bacillus subtilis spores. The choice of this biological systems is related to the fact that a number of reasons support the use of spores as a mucosal delivery system. The remarkable and well-documented resistance of spores to various environmental and toxic effects make them clear potentials as a novel, surface-display system. Our experimental outcomes demonstrate that TERS is able to provide a nano-scale chemical imaging of spore surface. Moreover, we demonstrate that TERS allows differentiation between wilde-type spore and genetically modified strains. These results hold promise for the characterization and optimization of spore surface for drug-delivery applications.
Cell-based biosensors rely on the detection and identification of single cells as well as monitoring of changes induced by interaction with drugs and/or toxic agents. Raman spectroscopy is a powerful tool to reach this goal, being non-destructive analytical technique, allowing also measurements of samples in aqueous environment. In addition, micro-Raman measurements do not require preliminary sample preparation (as in fluorescence spectroscopy), show a finger-print spectral response, allow a spatial resolution below typical cell sizes, and are relatively fast (few s or even less). All these properties make micro-Raman technique particularly promising for high-throughput on-line analysis integrated in lab-on-a-chip devices. Herein, we demonstrate some applications of Raman analysis in ophthalmology. In particular, we demonstrate that Raman analysis can provide useful information for the therapeutic treatment of keratitis caused by Acanthamoeba Castellanii (A.), an opportunistic protozoan that is widely distributed in the environment and is known to produce blinding keratitis and fatal encephalitis. In particular, by combining Raman analysis with Principal Component Analysis (PCA), we have demonstrated that is possible to distinguish between live and dead cells, enabling, therefore to establish the effectiveness of therapeutic strategies to vanquish the protozoa. As final step, we have analyzed the presence of biochemical differences in the conjunctival epithelial tissues of patients affected by keratitis with respect to healthy people. As a matter of facts, it is possible to speculate some biochemical alterations of the epithelial tissues, rendering more favorable the binding of the protozoan. The epithelial cells were obtained by impression cytology from eyes of both healthy and keratitis-affected individuals. All the samples were analyzed by Raman spectroscopy within a few hours from cells removal from eyes. The results of this analysis are discussed.
In this communication, we discuss the application of ordered, ultrahigh-density templates of nano-textured Ag-particles obtained by self-assembling of inorganic-containing polystyrene-block-poly(4-vinylpyridine) copolymer (PS-b-P4VP) micelles, for the spectroscopic surface-enhanced Raman imaging in-vitro of red blood cells (RBCs) and its capability to identify the vibrational fingerprint of the plasma membrane of the cell physisorbed to the SERS substrate. Hexagonal arrays of PS-b-P4VP micelles, with selective inclusion of Ag nanoparticles (NPs) in the polar core, prepared by in situ reduction of a suitable precursor, are obtained by polymer self-assembly upon fast solvent evaporation during spin coating on the supporting substrate. UV irradiation and/or plasma oxygen treatment remove the polymer matrix leaving immobilized nano-islands of Ag-NPs. Such a kind of SERS-active substrate consists of a reproducible and uniform twodimensional hexagonal array of silver clusters with a diameter ranging from 25 to 30 nm (single particles having typically diameters of 5 nm) and nano-island gap distances of the order of 5-8 nm on silicon and 15 nm on glass , while giving rise to high enhancement factors and addressing the issue of SERS reproducibility. The basic substrate supporting the plasmonic coating used in this work is either of silicon or glass. This last allows working in back scattering configuration permitting real time monitoring, via microscopy, of the RBCs on which Raman measurements are being carried out. The template is thus applied for surface-enhanced Raman analysis of the red blood cell (RBC) membrane in confocal micro-Raman configuration demonstrating to have SERS imaging potential thanks to the uniformity of the nano-textured substrate. The first experimental evidence of SERS imaging of a red blood cell membrane in-vitro is demonstrated.
Optical tweezers use the gradient force created by tightly focused single laser beam to trap dielectric microparticles. While this technique has been used for over 20 years to manipulate particles without mechanical contact, it is only recently htat accurate and quantitative photonic forces measurements have been considered. Moreover of great relevance has resulted the monitoring of Brownian motion of particles confined in optical traps since that provides precious information on local visco-elastic properties of the surrounding fluid. In this work we consider the still open question concerning the calibration of an optical tweezers which represents the key point for any absolute measurements. In particular, we discuss a novel method to calibrate a quadrant photodiode used as sensor position in the forward scattering scheme. The voltage signals provided by the qudrant photodiode are converted in length units by comparing them with the absolute bead positions measured by means of a calibrated CCD camera. Finally we briefly discuss how calibrated optical tweezers, combined with thermal analysis of the Brownian motion, are potentially of great relevance for microrheological studies of complex fluids.
Optical tweezers have become a widely used tool for manipulate microscopic objects. Combined with fast and high sensitive position detection techniques, they are suitable for microrheological measurements of viscous and viscoelastic media. Such measurements require the knowledge of absolute displacements of the trapped particles. Unfortunately position detection device need to be calibrated and several methods have been used and reported in literature. We report an accurate study of the behavior of the conversion factor in the case of the forward light scattering technique, based on an oil-immersion objective lens, which is commonly used in optical tweezers systems. We show how this fundamental parameter is strongly dependent on the distance of the trapped object from the coverslip surface.
We report an experimental study of the trapping efficiency of an optical tweezers system acting on spherical polystyrene particles in water solution. As force calibration method we used the viscous drag exerted by fluid flow. A parametric study of the efficiency of the trap was made as a function of bead diameter and objective NA. Optical tweezers were also characterized in terms of the optical potential well by measuring the displacement of trapped particles undergone to a viscous drag for fluid flow below the critical velocity. Trap stiffness was determined for particle size of 14.9 micrometers at different laser power.
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