Francesca Costantini

Nanostructure Based on Polymer Brushes for Efficient Heterogeneous Catalysis in Microreactors

PGMA polymer brushes are successfully grown on the inner wall of a microreactor to give a nanostructure. The oxirane groups of the brushes are used for the anchoring of a catalyst. The utility of the combination of catalyst-functionalized brushes and a microreactor is clearly demonstrated for the TBD-catalyzed Knoevenagel condensation reaction of benzaldehyde and malononitrile.

A Brush‐Gel/Metal‐Nanoparticle Hybrid Film as an Efficient Supported Catalyst in Glass Microreactors

A polymer-brush-based material was applied for the formation and in situ immobilization of silver and palladium nanoparticles, as a catalytic coating on the inner wall of glass microreactors. The brush film was grown directly on the microchannel interior by means of atom-transfer radical polymerization (ATRP), which allows control over the polymer film thickness and therefore permits the tuning of the number of nanoparticles formed on the channel walls. The wide applicability of the catalytic devices is demonstrated for the reduction of 4-nitrophenol and for the Heck reaction.

On-chip detection of multiple serum antibodies against epitopes of celiac disease by an array of amorphous silicon sensors

In this paper, we present the preliminary results of an ELISA-on-chip device, intended as a technological demonstrator of a novel analytical system suitable for the diagnosis and follow-up of celiac disease. The idea of the work is to combine an array of amorphous silicon photosensors with a pattern of a poly(2-hydroxyethyl methacrylate) polymer brush film, which acts as anchor for the immobilization of gliadin peptides containing the celiac disease epitopes. Recognition relies on a sandwich immunoassay between antibodies against the peptides and secondary antibodies marked with horseradish peroxidase to obtain a chemiluminescent signal. Detection is based on the measurement of photocurrent induced in the array of amorphous silicon photosensors by the chemiluminescent signal. An ad-hoc procedure has been developed in order to enable the fabrication of the photodiode array and the polymer brush pattern on the two sides of the same glass substrate ensuring the compatibility of the different technological steps. The sensitivity and the selectivity of the chip for multiplex immunoassays were demonstrated using two gliadin peptides (VEA and DEC). In particular, we found that the average amount of the bound HRP revealed by our analytical protocol is 3.5(±0.3) × 10−6 pg μm−2 and 0.85(±0.3) × 10−6 pg μm−2 for specific and non-specific interactions, respectively.

Targeted LipoCEST Contrast Agents for Magnetic Resonance Imaging: Alignment of Aspherical Liposomes on a Capillary Surface

Molecular imaging is likely to have a significant impact onhealthcare through the early detection of disease on a cellularand molecular level. Among the clinical imaging modalities,magnetic resonance imaging (MRI) offers a unique combi-nation of advantages including the recording of anatomicaland contrast-enhanced images with a high spatial resolution,while avoiding the use of ionizing radiation. The use of MRIfor imaging sparse molecular epitopes present on diseasedcells is hampered by its low sensitivity, which can potentiallybe overcome with new contrast-amplifying nanocarriers.

Glucose level determination with a multi-enzymatic cascade reaction in a functionalized glass chip

In this work we show the functionalization of the interior of microfluidic glass chips with poly(2-hydroxyethyl methacrylate) polymer brushes as anchors for co-immobilization of the enzymes glucose-oxidase and horseradish peroxidase. The formation of the brush layer and subsequent immobilization of these enzymes have been characterized on flat surfaces by atomic force microscopy and Fourier transform infrared spectroscopy, and studied inside glass chips by field emission scanning microscopy. Enzyme-functionalized glass chips have been applied for performing a multi-enzymatic cascade reaction for the fast (20 s) determination of glucose in human blood samples and the result is in excellent agreement with values obtained from the conventional hospital laboratory. The limit of detection of this bi-enzymatic method is 60 μM. With the advantages of high selectivity and reproducibility, this functionalization method can be used for improving the efficiency of glucose sensors.

On-Chip Diagnosis of Celiac Disease by an Amorphous Silicon Chemiluminescence Detector

A lab-on-chip for the diagnosis of celiac disease relying on the monitoring of patient-specific immune response to gliadin fractions has been developed. The detection is based on a chemiluminescent immunoenzymatic reaction that ensures high specificity and sensitivity. The chemiluminescent signal is monitored by hydrogenated amorphous silicon photosensors, fabricated on the same glass substrate hosting the biochemical recognition. The main challenge of the work has been the identification of the materials and the setup of the entire process that permitted the reliable fabrication of the device. Experiments performed with serum samples of rabbit immunized towards an epitope show a good specificity of the proposed technique, proving the feasibility of an integrated device for the patient-specific profiling.

Dimensional scale effects on surface enhanced Raman scattering efficiency of self-assembled silver nanoparticle clusters

A study of the Surface Enhanced Raman Scattering (SERS) from micrometric metallic nanoparticle aggregates is presented. The sample is obtained from the self-assembly on glass slides of micro-clusters of silver nanoparticles (60 and 100 nm diameter), functionalized with the organic molecule 4-aminothiophenol in water solution. For nanoparticle clusters at the micron scale, a maximum enhancement factor of 109 is estimated from the SERS over the Raman intensity ratio normalized to the single molecule contribution. Atomic force microscopy, correlated to spatially resolved Raman measurements, allows highlighting the connection between morphology and efficiency of the plasmonic system. The correlation between geometric features and SERS response of the metallic structures reveals a linear trend of the cluster maximum scattered intensity as a function of the surface area of the aggregate. On given clusters, the intensity turns out to be also influenced by the number of stacking planes of the aggregate, thus sugg...

Multifunctional System-on-Glass for Lab-on-Chip applications.

Lab-on-Chip are miniaturized systems able to perform biomolecular analysis in shorter time and with lower reagent consumption than a standard laboratory. Their miniaturization interferes with the multiple functions that the biochemical procedures require. In order to address this issue, our paper presents, for the first time, the integration on a single glass substrate of different thin film technologies in order to develop a multifunctional platform suitable for on-chip thermal treatments and on-chip detection of biomolecules. The proposed System on-Glass hosts thin metal films acting as heating sources; hydrogenated amorphous silicon diodes acting both as temperature sensors to monitor the temperature distribution and photosensors for the on-chip detection and a ground plane ensuring that the heater operation does not affect the photodiode currents. The sequence of the technological steps, the deposition temperatures of the thin films and the parameters of the photolithographic processes have been optimized in order to overcome all the issues of the technological integration. The device has been designed, fabricated and tested for the implementation of DNA amplification through the Polymerase Chain Reaction (PCR) with thermal cycling among three different temperatures on a single site. The glass has been connected to an electronic system that drives the heaters and controls the temperature and light sensors. It has been optically and thermally coupled with another glass hosting a microfluidic network made in polydimethylsiloxane that includes thermally actuated microvalves and a PCR process chamber. The successful DNA amplification has been verified off-chip by using a standard fluorometer.

Folate-based single cell screening using surface enhanced Raman microimaging

Recent progress in nanotechnology and its application to biomedical settings have generated great advantages in dealing with early cancer diagnosis. The identification of the specific properties of cancer cells, such as the expression of particular plasma membrane molecular receptors, has become crucial in revealing the presence and in assessing the stage of development of the disease. Here we report a single cell screening approach based on Surface Enhanced Raman Scattering (SERS) microimaging. We fabricated a SERS-labelled nanovector based on the biofunctionalization of gold nanoparticles with folic acid. After treating the cells with the nanovector, we were able to distinguish three different cell populations from different cell lines (cancer HeLa and PC-3, and normal HaCaT lines), suitably chosen for their different expressions of folate binding proteins. The nanovector, indeed, binds much more efficiently on cancer cell lines than on normal ones, resulting in a higher SERS signal measured on cancer cells. These results pave the way for applications in single cell diagnostics and, potentially, in theranostics.

Integrated chemiluminescence-based lab-on-chip for detection of life markers in extraterrestrial environments

The detection of life markers is a high priority task in the exploration of the Solar System. Biochips performing in-situ multiplex immunoassays are a very promising approach alternative to gas chromatography coupled with mass spectrometry. As part of the PLEIADES project, we present the development of a chemiluminescence-based, highly integrated analytical platform for the detection of biomarkers outside of the Earth. The PLEIADES device goes beyond the current lab-on-chip approaches that still require bulky external instrumentation for their operation. It exploits an autonomous capillary force-driven microfluidic network, an array of thin-film hydrogenated amorphous silicon photosensors, and chemiluminescence bioassays to provide highly sensitive analyte detection in a very simple and compact configuration. Adenosine triphosphate was selected as the target life marker. Three bioassay formats have been developed, namely (a) a bioluminescence assay exploiting a luciferase mutant with enhanced thermal and pH stability and (b and c) binding assays exploiting antibodies or functional nucleic acids (aptamers) as biospecific recognition elements and peroxidase or DNAzymes as chemiluminescence reporters. Preliminary results, showing limits of detection in the nanomolar range, confirm the validity of the proposed approach.