Elettra Synchrotron Light Laboratory

Trieste, Italy

Elettra Synchrotron Light Laboratory

Trieste, Italy

Time filter

Source Type

Mitri E.,CNR Institute of Materials | Mitri E.,University of Trieste | Birarda G.,Lawrence Berkeley National Laboratory | Vaccari L.,Elettra Synchrotron Light Laboratory | And 4 more authors.
Lab on a Chip - Miniaturisation for Chemistry and Biology | Year: 2014

Here we present a new bonding protocol for SU-8 negative tone photoresist that exploits the chemical modifications induced in the resin by exposure to 254 nm (UVC) light. Fourier Transform Infrared microspectroscopy (μ-FTIR) was used to carry out a thorough study on the chemical processes and modifications occurring within the epoxy resin by exposure to 365 nm and 254 nm light. In particular, we established that UVC light promotes the opening of the epoxy rings bypassing the post-exposure bake. The possibility to promote a further activation of the resin, already patterned with standard UV lithography, was exploited to produce closed microfluidic devices. Specifically, we were able to fabricate fluidic chips, characterized by broadband transparency from mid-IR to UV and long term stability in continuous flow conditions. CaF2 was used as substrate, coated by sputtering with a nanometric silicon film, in order to make surface properties of this material more suitable for standard fabrication processes with respect to the original substrate. The fabricated microfluidic chips were used to study by μ-FTIR the biochemical response of live breast cancer MCF-7 cells to osmotic stress and their subsequent lysis induced by the injection of deionized water in the device. μ-FTIR analyses detected fast changes in protein, lipid and nucleic acid content as well as cytosol acidification. This journal is © 2014 The Royal Society of Chemistry.


Mitri E.,CNR Institute of Materials | Mitri E.,University of Trieste | Pozzato A.,CNR Institute of Materials | Coceano G.,CNR Institute of Materials | And 6 more authors.
Microelectronic Engineering | Year: 2013

In this contribution we present the first example of a microfluidic chip based on BaF2 for Infrared Microspectroscopy (IRMS) of living cells. The advantage in using barium fluoride as platform relies on its high IR transparency, especially in the spectral region below 1300 cm-1, where the absorption bands of nucleic acids and carbohydrates are located. Barium fluoride is slightly soluble in water (0.12 g/100 g water) and it is potentially harmful for living cells. To overcome these problems, here we exploit an approach whose feasibility has been demonstrated previously on CaF2: the surface modification obtained by sputtering a thin Si layer on the surface. The Surface Modified Microfluidic Devices (SM-MD) hence obtained not only solve the BaF2 drawbacks, but also provide a silicon-like substrate fully compatible with standard micro-fabrication processes. These potentialities are here further explored in the direction of chemical or topographical nano-patterning of the silicon-like surface. The silicon thin layer was structured in the shape of 300 nm wide grooves (500 nm pitch) with a thickness of 35 nm by using standard NIL and etching processes; chemical patterning was achieved by exploiting silane chemistry. Finally, we tested the performances of these devices at SISSI beamline@Elettra, collecting IR spectra of single MDA-MB-231 living cells maintained either in physiological solution or complete medium. A comparison of the spectra of a single cell obtained in BaF2 and CaF2 MDs is reported. © 2013 Elsevier B.V. All rights reserved.


D'amico F.,Elettra Synchrotron Light Laboratory | Saito M.,Elettra Synchrotron Light Laboratory | Bencivenga F.,Elettra Synchrotron Light Laboratory | Marsi M.,University Paris - Sud | And 10 more authors.
Nuclear Instruments and Methods in Physics Research, Section A: Accelerators, Spectrometers, Detectors and Associated Equipment | Year: 2013

We present a newly developed resonant Raman scattering instrument working in the UV spectral range. This set-up, which exploits the UV synchrotron radiation source available at Elettra (Trieste, Italy), results in an innovative spectroscopic facility to be used for addressing a large array of open problems, ranging from the electronic properties of nanostructures and strongly correlated materials to biochemistry. © 2012 Elsevier B.V.


Vaccari L.,Elettra Synchrotron Light Laboratory | Birarda G.,Elettra Synchrotron Light Laboratory | Birarda G.,CNR Institute of Materials | Businaro L.,CNR Institute for Photonics and Nanotechnologies | And 2 more authors.
Analytical Chemistry | Year: 2012

Until nowadays most infrared microspectroscopy (IRMS) experiments on biological specimens (i.e., tissues or cells) have been routinely carried out on fixed or dried samples in order to circumvent water absorption problems. In this paper, we demonstrate the possibility to widen the range of in-vitro IRMS experiments to vibrational analysis of live cellular samples, thanks to the development of novel biocompatible IR-visible transparent microfluidic devices (MD). In order to highlight the biological relevance of IRMS in MD (MD-IRMS), we performed a systematic exploration of the biochemical alterations induced by different fixation protocols, ethanol 70% and formaldehyde solution 4%, as well as air-drying on U937 leukemic monocytes by comparing their IR vibrational features with the live U937 counterpart. Both fixation and air-drying procedures affected lipid composition and order as well as protein structure at a different extent while they both induced structural alterations in nucleic acids. Therefore, only IRMS of live cells can provide reliable information on both DNA and RNA structure and on their cellular dynamic. In summary, we show that MD-IRMS of live cells is feasible, reliable, and biologically relevant to be recognized as a label-free cell-based assay. © 2012 American Chemical Society.


Flower K.R.,University of Manchester | Khalifa I.,University of Manchester | Bassan P.,University of Manchester | Demoulin D.,University of Manchester | And 6 more authors.
Analyst | Year: 2011

Recently a new di-gold(i) organometallic complex [1,3-(Ph 3PAu)2-C6H4] (KF0101) has been synthesised and found to exhibit cytotoxic activity in vitro. Subsequently it has been demonstrated that KF0101 shows little or no cross-resistance against a number of the cisplatin resistant ovarian cancer cell lines in vitro suggesting a different mode of action for the drug. In this study, syncrotron radiation infrared microspectroscopy (SR-IRMS) has been used on drug treated single A2780 cells in order to determine if this different mode of action can be identified spectroscopically. The aim of the study was to establish: (i) if single cell SR-IRMS could be used to give insight into the cellular response on treatment with different cytotoxic agents relative to non-treated cells (control) and (ii) that if the cytotoxic drugs elicit a different biochemical response these responses could be distinguished from each other. The most striking features obtained after Principal Componants Analysis (PCA) of Resonant Mie Scattering (RMieS) corrected single cell spectra of drug treated ovarian A2780 cells are: (i) The spectra obtained for the control are quite heterogeneous and several hundred spectra are required to adequately define the nature of the control; (ii) after drug treatment at the IC50 level for 24 h with cisplatin, KF0101, methotrexate, paclitaxel or 5-fluorouracil the cell spectra, as represented on a PCA scores plot, generally concentrate in certain well defined areas of the control, there are however a small number of spectra that fall outside of the area defined by the control; and (iii) a differentiation between cell spectra obtained on treatment with different drugs is observed which fits well with different in vitro cell culture behaviour and a flow cytometry cell cycle analysis of the contol and drug treated cells. Inspection of the loading plots shows that PC1 is essentially the same for all plots and reflects changes in cell biochemistry related to the cell cycle. PC2, however, on comparison of the control versus cisplatin or cisplatin versus KF0101 is indicative of differences induced by drug treatment and has been termed as cell cycle-plus behaviour. These data are shown to be consistent with that obtained using bench-top IRMS by averaging a number of single cell spectra and carrying out a PCA, but SR-IRMS offers more insight into how the drug is affecting the cell population. More importantly, this approach enables the influence of the cell cycle on both the control and drug treated samples to be taken into consideration when evaluating the drug-cell interaction. © 2011 The Royal Society of Chemistry.


Birarda G.,Elettra Synchrotron Light Laboratory | Birarda G.,CNR Institute of Neuroscience | Birarda G.,University of Trieste | Grenci G.,CNR Institute of Neuroscience | And 6 more authors.
Vibrational Spectroscopy | Year: 2010

First experiments demonstrating the suitability of novel microfabricated fluidic devices for measuring living cells in physiological environment by synchrotron radiation (SR) Fourier Transform Infrared microspectroscopy (μ-FTIR) are presented. The devices were fabricated on CaF2 windows, using the photoresist XARP 3100/10 to define the liquid cell lay-out. Therefore, the sample holder is transparent to both visible and infrared light, robust, completely recyclable and with a precise spacing. Using prototype devices of thicknesses 9, 5 and 3 μm, we studied the response of the U937 monocytic cell line to mechanical compression. The temporal evolution of the FTIR spectra, characteristic for the status of living cells, was used to monitor the cellular system stability in time. Protein biosynthesis and lipid metabolism alterations upon deformation have been identified by monitoring specific cell band ratios such as amide I to amide II, amide I to lipids, methylene to methyl and asymmetric to symmetric stretching of phosphates. Taking advantage of the high brilliance of the synchrotron radiation, chemical maps of monocyte cells were collected, demonstrating the versatility of the device. © 2010 Elsevier B.V. All rights reserved.


Birarda G.,Elettra Synchrotron Light Laboratory | Birarda G.,CNR Institute of Neuroscience | Birarda G.,University of Trieste | Grenci G.,CNR Institute of Neuroscience | And 4 more authors.
Microelectronic Engineering | Year: 2010

Here we present the optimization of fabrication steps for realizing an infrared-visible microfluidic chip to study single-living cell behaviour in physiological environment by synchrotron radiation FTIR microspectroscopy. We optimized subtractive and additive lithographic processes on CaF2 substrate, employing X-ARP 3100/10 photoresist both as etching-mask and for the device fabrication. Using prototype microfabricated liquid cells 9 and 5 μm thick, we measured the response of small groups of THP1 monocytic cells to mechanical compression and chemical stimulation with fMLP using conventional IR globar source, aiming to evaluate biochemical rearrangements of leukocytes during the capillary circulation or recruitment processes. Stimulated monocytes have spectral features recognizable, differentiating them from un-stimulated, especially affecting the spectral region 1280-1000 cm-1, characteristic of nucleic acids and carbohydrates, and specific band ratios, such as proteins on lipids and methylene on methyl. Spectra variations have been correlated with biochemical events such as transcription, synthesis of new-proteins and variations in membrane fluidity. © 2009 Elsevier B.V. All rights reserved.


Gioacchini G.,Center for Reproductive Health | Gioacchini G.,Marche Polytechnic University | Giorgini E.,Marche Polytechnic University | Vaccari L.,Elettra Synchrotron Light Laboratory | And 5 more authors.
Fertility and Sterility | Year: 2014

Objective To characterize from a vibrational point of view the alterations caused by aging on human oocytes. Design Reproductive biology. Setting Private assisted reproductive technology clinic, synchrotron beam line, and university infrared laboratory. Patient(s) Twenty women of different ages (30 ± 2 and 39 ± 2 years) selected on the basis of detailed inclusion criteria and submitted to controlled ovarian stimulation according to a specific protocol. Intervention(s) Collection of 68 supernumerary oocytes that were not used during the IVF cycle from the above cited consenting patients. Main Outcome Measure(s) Focal Plane Array Fourier transform infrared (FTIR) analysis of human oocytes. Result(s) Specific spectral differences were highlighted in the two experimental groups of oocytes. In particular, in oocytes of 39-year-old women, the occurrence of peroxidative processes and a decrease in the amount of carbohydrates were observed, together with alterations in the phospholipid membrane, proteic pattern, and nucleic acids content. Conclusion(s) For the first time, FTIR spectroscopy was applied to human oocytes, leading to strong evidence of damage from aging in the gametes of mature women, which could be related to a decline in reproductive function. All the information obtained may be considered useful to improve the scientific knowledge on human reproduction and to exploit new strategies for detecting oocyte aging. © 2014 by American Society for Reproductive Medicine.


Loutherback K.,Lawrence Berkeley National Laboratory | Birarda G.,Lawrence Berkeley National Laboratory | Birarda G.,Elettra Synchrotron Light Laboratory | Chen L.,Lawrence Berkeley National Laboratory | Holman H.-Y.N.,Lawrence Berkeley National Laboratory
Protein and Peptide Letters | Year: 2016

A long-standing desire in biological and biomedical sciences is to be able to probe cellular chemistry as biological processes are happening inside living cells. Synchrotron radiation-based Fourier transform infrared (SR-FTIR) spectral microscopy is a label-free and nondestructive analytical technique that can provide spatiotemporal distributions and relative abundances of biomolecules of a specimen by their characteristic vibrational modes. Despite great progress in recent years, SR-FTIR imaging of living biological systems remains challenging because of the demanding requirements on environmental control and strong infrared absorption of water. To meet this challenge, microfluidic devices have emerged as a method to control the water thickness while providing a hospitable environment to measure cellular processes and responses over many hours or days. This paper will provide an overview of microfluidic device development for SR-FTIR imaging of living biological systems, provide contrast between the various techniques including closed and open-channel designs, and discuss future directions of development within this area. Even as the fundamental science and technological demonstrations develop, other ongoing issues must be addressed; for example, choosing applications whose experimental requirements closely match device capabilities, and developing strategies to efficiently complete the cycle of development. These will require imagination, ingenuity and collaboration. © 2016 Bentham Science Publishers.


PubMed | Elettra Synchrotron Light Laboratory
Type: Journal Article | Journal: Analytical chemistry | Year: 2012

Until nowadays most infrared microspectroscopy (IRMS) experiments on biological specimens (i.e., tissues or cells) have been routinely carried out on fixed or dried samples in order to circumvent water absorption problems. In this paper, we demonstrate the possibility to widen the range of in-vitro IRMS experiments to vibrational analysis of live cellular samples, thanks to the development of novel biocompatible IR-visible transparent microfluidic devices (MD). In order to highlight the biological relevance of IRMS in MD (MD-IRMS), we performed a systematic exploration of the biochemical alterations induced by different fixation protocols, ethanol 70% and formaldehyde solution 4%, as well as air-drying on U937 leukemic monocytes by comparing their IR vibrational features with the live U937 counterpart. Both fixation and air-drying procedures affected lipid composition and order as well as protein structure at a different extent while they both induced structural alterations in nucleic acids. Therefore, only IRMS of live cells can provide reliable information on both DNA and RNA structure and on their cellular dynamic. In summary, we show that MD-IRMS of live cells is feasible, reliable, and biologically relevant to be recognized as a label-free cell-based assay.

Loading Elettra Synchrotron Light Laboratory collaborators
Loading Elettra Synchrotron Light Laboratory collaborators