257 research outputs found
Raman spectroscopy determination of the mineral characteristics of microcalcifications in breast cancer: a way towards an improved screening approach
No full textDEVELOPMENT OF A NOVEL NANOSENSOR FOR THE STUDY OF BIOMOLECULAR INTERACTIONS
No full textThe evaluation and quantification of molecular interactions is of paramount importance in modern biology and molecular medicine. Therefore, there is a continuous exploration for new methodologies capable to detect and to measure binding affinities during reversible
molecular interactions. This work is devoted to explore a new tool based on the high
sensitivity that the measurement of the scattered light intensity offers when the binding
occurs on the surface of index-matched colloids.
Static light scattering is not a traditional technique to study molecular association because
the binding of insulated ligands and receptors in dilute solutions produce negligible
increment of the scattered light, while mesoscopic particles hosting multiple receptors or
ligands, including real bacteria, typically scatter too much light compared with the
contributions due to molecular adhesion on their surface. This difficulty can be overcome
by supporting the receptors on nano-scale latex spheres whose refractive index closely
matches the one of water.
As “Phantom Nanoparticles” (PNPs), we have used highly hydrophobic monodisperse
spherical fluoroelastomer colloids, with radius R = 39 Å} 1 nm, and whose refractive index,
at our working temperature (30ÅãC) and wavelength (633 nm), is np0 = 1.3248 (under the
same conditions the refractive index of water is nW = 1.3319). Surfactants added to a PNP
dispersion readily adsorb on their hydrophobic surfaces, generating a self-assembled
monolayer which can be easily equipped with molecular hydrophilic end groups of various
kind, including well-known receptors and/or ligands.
This label-free method has been assessed through the precise determination of the binding
constant of the antibiotic vancomycin with the tripeptide L-Lys-D-Ala-D-Ala and of the
vancomycin dimerization constant. We have enlightened the role of bidentate effect and
molecular hindrance in the activity of this glycopeptide.
After this success first result, an accurate determination of the optimal properties of
nanoparticles employed has been performed by comparative experiments and through
theoretical evaluation (CHAPETR 3). The effects of size, refractive index, electric charge,
and dilution on the reliability and accuracy of the method has been evaluated. Quite
surprisingly, perfect index matching and minimal size (i.e., maximum surface), which is
almost attained in one of the colloids here employed, do not represent the ideal conditions.
Rather, we show that a nanoparticle radius of 100 nm and a refractive index slightly below
that of water yields the best signal/background amplitude. We also show that repulsive
interactions can lead to artifacts in the adsorption isotherm, thus indicating that
electrostatic stabilization should be kept at a minimum.
Successively, the particles, already optically phantom, have also been made biologically
“invisible” through PEG coating and decorated by interacting proteins, thus providing a
mean to investigate the biological properties of proteins (CHAPETR 4). Avidin decorated
Phantom Nanoparticles have been prepared and were employed to detect interactions
between different kinds of biotinylated proteins. Using this approach, biotinylated protein
A was anchored on the surface of the nanoparticles, and were exploited as a functional
probe for the rapid, quantitative, picomolar detection of human IgG antibodies.
We used Phantom Nanoparticles to evaluate substrate recognition by Streptomyces PMF
Phospholipase D inactivated mutants (CHAPETR 5). The use of this specific technique
seems to have some peculiar advantages over other methods in the case of phospholipidsacting
enzymes. In fact, the substrate (or a substrate analog) can be organized onto the
surface of Phantom Nanoparticles at a desired concentration with optimal display of the
polar head group, while the hydrophobic chains result packed into the surfactant
monolayer, thus limiting the occurrence of non-specific interactions.
The last part of this work is focused on an attempt toward the stable functionalization of
Phantom Nanoparticles (CHAPETR 6). Diacetylene surfactants spontaneously adsorb on the
nanoparticles and then they are polymerized by exposure to UV light. So far, the stability
of the amphiphilic coating around the nanoparticle solely depended on weak hydrophobic
interactions. The attachment of the polymer to the particle surface, because of the
numerous contact points, is highly stable and can be improved further by crosslinking of
the polymer shell. Huns generating a quantifiable number of functional groups suitable for
covalent receptor anchorage.
All these observations, demonstrate the feasibility of this new technique, which makes it
possible to easily generate different synthetic receptors, and highlight this technique as a
versatile novel method to study, both qualitatively and quantitatively, of molecular
recognition processes.
The work described in this thesis is partially published in:
Morasso C., Bellini T., Monti D., Bassi M., Prosperi D., Riva S.:
Dispersed phantom scatterer technique reveals subtle differences in
substrate recognition by phospholipase D inactive mutant;
ChemBioChem. Submitted, currently under review
Prosperi D., Morasso C., Tortora P., Monti D., Bellini T.: Avidin decorated
core-shell nanoparticles for biorecognition studies by elastic light
scattering; ChemBioChem. 2007 (8): 1021-1028 (Impact factor: 3,446).
Prosperi D., Morasso C., Mantegazza F., Buscaglia M., Houg L., Bellini
T.: Phantom nanoparticles as probes of biomolecular interaction; Small.
2006 (8-9):1060-1067 (Impact factor: 6,408)
A new look at posturographic analysis in the clinical context: sway-density vs. other parameterization techniques.
No full text
HMM-based anomaly interpretation for intelligent robots in Industry 4.0
No full textWe apply an anomaly detection method based on Hidden Markov Models and Hellinger distance to a Kairos mobile robot operating in the ICE lab, a research laboratory for Industry 4.0. Two main contributions are proposed: i) a decomposition of the Hellinger distance which allows to identify the causes of anomalous behaviours detected, ii) a graphical user interface that synchronously shows the robot movements in a map and the evolution of the Hellinger distance components, allowing a quick investigation of the causes of the detected anomalies. The tools are applied to a real-world dataset allowing to discover that an anomalous movement of the Kairos robot is caused by a wrong reading of the lidar from a window in the environment
Molecular Approach to Cutaneous Squamous Cell Carcinoma: From Pathways to Therapy
No full textCutaneous squamous cell carcinoma (cSCC) represents the second most frequent skin cancer, recently showing a rapid increase in incidence worldwide, with around >1 million cases/year in the United States and 2500 deaths [...
Abnormal sensorimotor control, but intact force field adaptation, in multiple sclerosis subjects with no clinical disability
No full textIn MS subjects with no clinical disability, we assessed sensorimotor organization and their ability to adapt to an unfamiliar dynamical environment. Eleven MS subjects performed reaching movements while a robot generated a speed-dependent force field. Control and adaptation performance were compared with that of an equal number of control subjects. During a familiarization phase, when the robot generated no forces, the movements of MS subjects were more curved, displayed greater and more variable directional errors and a longer deceleration phase. During the force field phase, both MS and control subjects gradually learned to predict the robot-generated forces. The rates of adaptation were similar, but MS subjects showed a greater variability in responding to the force field. These results suggest that MS subjects have a preserved capability of learning to predict the effects of the forces, but make greater errors when actually using such predictions to generate movements. Inaccurate motor commands are then compensated later in the movement through an extra amount of sensory-based corrections. This indicates that early in the disease MS subjects have intact adaptive capabilities, but impaired movement execution
A haptic robot reveals the adaptation capability of individuals with Multiple Sclerosis
No full textA prerequisite for rehabilitation is that patients preserve their ability to adapt to novel dynamic environments, an ability that has been associated with the cerebellar system. In this study, we use a robot manipulandum to assess the ability of multiple sclerosis (MS) sub- jects in the early phase of the disease to adapt to a speed-dependent force field. Their performance is compared with an equal number of age-matched controls. We found that MS subjects display subtle in- coordination problems but do not significantly differ from controls in their ability to adapt to the force field. These findings are discussed in terms of the possible benefits that MS subjects might receive from robot-assisted therapy that is specifically aimed at impaired visuomo- tor coordination
Avidin decorated core–shell nanoparticles for biorecognition studies by elastic light scattering
No full textIn this paper, a straightforward method based on elastic light scattering is shown to provide a sensitive and reliable tool for the quantitative determination of protein-ligand interactions that occur at the surface of suitably designed core-shell nanoparticles. The assay makes use of monodisperse nanocolloids that have minimal optical contrast with the aqueous environment. By properly coating the particles with avidin and oligo(ethylene glycol)-based amphiphiles, we developed a hybrid system that combines the availability of standard ligands with the necessary bioinvisibility towards the accidental adsorption of nonspecific macromolecules. This probe was employed to detect interactions between different kinds of biotinylated proteins, and it revealed high specificity and affinities in the low nanomolar range. In particular, we obtained an efficient avidin anchorage of biotinylated protein A on the surface of the nanoparticles, which we exploited as a functional probe for the rapid, quantitative, picomolar detection of human IgG antibodies. Overall, these light-scattering-based nanosensors appear as a simple and highly informative tool for proteomics studies
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