Fulvia Sinatra

XPS and AFM Characterization of the Enzyme Glucose Oxidase Immobilized on SiO2 Surfaces

A process to immobilize the enzyme glucose oxidase on SiO2 surfaces for the realization of integrated microbiosensors was developed. The sample characterization was performed by monitoring, step by step, oxide activation, silanization, linker molecule (glutaraldehyde) deposition, and enzyme immobilization by means of XPS, AFM, and contact angle measurements. The control of the environment during the procedure, to prevent silane polymerization, and the use of oxide activation to obtain a uniform enzyme layer are issues of crucial importance. The correct protocol application gives a uniform layer of the linker molecule and the maximum sample surface coverage. This result is fundamental for maximizing the enzyme bonding sites on the sample surface and achieving the maximum surface coverage. Thin SiO2 layers thermally grown on a Si substrate were used. The XPS Si 2p signal of the substrate was monitored during immobilization. Such a signal is not completely shielded by the thin oxide layer and it is fully suppressed after the completion of the whole protocol. A power spectral density analysis on the AFM measurements showed the crucial role of both the oxide activation and the intermediate steps (silanization and linker molecule deposition) to obtain uniform immobilized enzyme coverage. Finally, enzymatic activity measurements confirmed the suitability of the optimized protocol.

Modification of graphene oxide by laser irradiation: a new route to enhance antibacterial activity.

The antibacterial activity and possible toxicity of graphene oxide and laser-irradiated graphene oxide (iGO) were investigated. Antibacterial activity was tested on Escherichia coli and shown to be higher for GO irradiated for at least three hours, which seems to be correlated to the resulting morphology of laser-treated GO and independent of the kind and amount of oxygen functionalities. X-ray photoelectron spectroscopy, Raman spectroscopy, dynamic light scattering and scanning electron microscopy (SEM) show a reduction of the GO flakes size after visible laser irradiation, preserving considerable oxygen content and degree of hydrophilicity. SEM images of the bacteria after the exposure to the iGO flakes confirm membrane damage after interaction with the laser-modified morphology of GO. In addition, a fish embryo toxicity test on zebrafish displayed that neither mortality nor sublethal effects were caused by the different iGO solutions, even when the concentration was increased up to four times higher than the one effective in reducing the bacteria survival. The antibacterial properties and the absence of toxicity make the visible laser irradiation of GO a promising option for water purification applications.

Ultrastructural features and ICSI treatment of severe teratozoospermia: report of two human cases of globozoospermia

Abnormal sperm morphology is associated with male infertility. We describe two human cases of globozoospermia (round-headed spermatozoa) together with fine diagnosis and proposed treatment. Scanning and transmission electron microscopy (SEM and TEM) were performed to identify the ultrastructural features. Female partners underwent ovarian hyperstimulation and intracytoplasmic sperm injection (ICSI). Fertilized oocytes were transferred 36 h later. One couple had a healthy live-birth. Ultrastructural analysis may help to diagnose sperm morphology and identify those which will respond to treatment.

Feasibility Studies on Si-Based Biosensors

The aim of this paper is to summarize the efforts carried out so far in the fabrication of Si-based biosensors by a team of researchers in Catania, Italy. This work was born as a collaboration between the Catania section of the Microelectronic and Microsystem Institute (IMM) of the CNR, the Surfaces and Interfaces laboratory (SUPERLAB) of the Consorzio Catania Ricerche and two departments at the University of Catania: the Biomedical Science and the Biological Chemistry and Molecular Biology Departments. The first goal of our study was the definition and optimization of an immobilization protocol capable of bonding the biological sensing element on a Si-based surface via covalent chemical bonds. We chose SiO2 as the anchoring surface due to its biocompatibility and extensive presence in microelectronic devices. The immobilization protocol was tested and optimized, introducing a new step, oxide activation, using techniques compatible with microelectronic processing. The importance of the added step is described by the experimental results. We also tested different biological molecule concentrations in the immobilization solutions and the effects on the immobilized layer. Finally a MOS-like structure was designed and fabricated to test an electrical transduction mechanism. The results obtained so far and the possible evolution of the research field are described in this review paper.

Resveratrol and propolis extract: an insight into the morphological and molecular changes induced in DU145 cells.

In the Western world cancer is the second leading cause of mortality, and prostate carcinoma represents in men the second most important type of cancer-causing death. We have already shown that resveratrol (200 microM) triggers in DU145, an androgen-resistant prostate cancer cell line, a necrotic-like cell death, while propolis ethanolic extract (100 microg/ml) causes an apoptotic-like cell demise. The present research is aimed to better elucidate the molecular mechanisms activated by the two micronutrients. Vinorelbine bitartrate, a drug widely used in prostate cancer therapy, was utilized as a reference drug, because it is known to induce apoptosis. The combined treatments between the micronutrients and vinorelbine have been studied to test a possible vinorelbine dose reduction, avoiding its side effects without altering its cytotoxic action. In this investigation SEM and TEM analyses were performed to examine the morphological modifications induced; our observations confirmed necrotic cell features after treatment with resveratrol, and apoptotic modifications after propolis. We also measured cell cycle progression to study a correlation with p21 and p53, two well-known cell cycle checkpoints. The levels of HSP27 and HSP70, two chaperones also exerting antioxidant/antiapoptotic functions, were been also analyzed. Our data indicate that the two micronutrients modulate cell cycle distribution, increasing p53 levels, without the induced HSPs being able to rescue DU145 from death. The results presented suggest chemotherapy based on resveratrol and propolis, alone or in combination with vinorelbine, as a potential useful tool for prostate cancer therapy; the increase in cell cycle control and the modulation of HSPs expression reinforce this suggestion.

Glucose oxidase characterization for the fabrication of hybrid microelectronic devices

We studied the enzyme glucose oxidase (GOx) immobilization on silicon oxide surfaces. In particular, we optimized the immobilization protocol and verified that it fulfills both requirements of enzyme preservation (measured by enzymatic activity) and VLSI compatibility. The immobilization consists of four steps: oxide activation, silanization, linker molecule deposition and GOx immobilization. It is crucial to form an uniform linker layer on the sample surface in order to maximize the sites available for enzyme bonding and achieving the best enzyme deposition. In this study, utilizing glutaraldehyde as bifunctional reagent, we monitored its uniformity on the surface through X-ray Photoelectron Spectroscopy (XPS). Once optimized, the same protocol was used to anchor the enzyme in a porous silicon dioxide matrix. Gold labeled GOx molecules were monitored by electron diffraction X-ray (EDX) measurements coupled with scanning electron microscopy (SEM). The enzymatic activity was also monitored to confirm the goodness of the proposed immobilization method. Finally, the electrical characterization of MOS capacitors, showing a shift of about 1 V in the flat band voltage, demonstrated the possibility to use this approach for electrical detection.

New method for the detection of enzyme immobilized on Si-based glucose Biosensors

Glucose oxidase molecular layers immobilized on planar and porous SiO2 substrates were characterized with techniques commonly used in microelectronics technology. Both planar and porous SiO2 were used as inorganic platforms for the enzyme bonding. The immobilization protocol on planar SiO2 was characterized using contact angle, atomic force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS), while energy dispersive X-ray analysis (EDX) coupled with scanning electron microscopy (SEM) were used on porous SiO2 samples. A conclusive confirmation of the enzyme presence in the SiO2 matrix was obtained by gold nano particles labeling. This last measurement technique could be implemented to determine the enzyme concentration in the porous SiO2 layer.

An investigation into the physico-chemical conditions of the coastal waters around the Gulf of Catania

Abstract The results are given of sea water sampling at 49 different stations around the Gulf of Catania. The main physico-chemical parameters are reported (temperature, pH, dissolved oxygen, salinity, BOD) as well as the concentrations of some nutritive salts (nitrite, nitrate, phosphate).

Bone morphogenic protein BMP7 induces adipocyte differentiation and uncoupling protein UCP1 expression in human bone marrow mesenchymal stem cells

In this work human bone marrow mesenchymal stem cells (hMSCs) have been differentiated into brown adipocytes by the bone morphogenic protein BMP7, and the morphological and biochemical changes of hMSCs occurring during the differentiation towards brown adipocytes have been investigated. After differentiation, the cells display the presence of numerous vacuoles full of lipids; staining with Oil Red reveals the typical multilocular organization of fat, and a compact network of actin microfilaments surrounding fat vacuoles appears. Uncoupling protein 1 (UCP1), a mitochondrial protein characteristic of the brown adipose tissue, only visible by Western blotting in hMSCs, exhibits a marked increase after the second cycle of adipogenic induction and even more after the fourth cycle, while the mitochondria change their localization from scattered throughout the cytoplasm to mainly near the fat vacuoles. In differentiated brown adipocytes mitochondria close to the vacuoles display a low membrane potential most likely due to the abundant presence of UCP1. Hopefully the additional knowledge about the role of UCP1 in hMSCs and brown adipocytes will be useful in developing strategies for the treatment of several diseases, such as obesity and type 2 diabetes.

Egg envelopes of Armadillidium vulgare (Latreille, 1804) (Crustacea, Isopoda Oniscidea): Ultrastructure and lectins binding

The ultrastructural study carried out on (a) oocytes of Armadillidium vulgare during vitellogenesis, (b) mature eggs taken from the ovaries during the parturial moult of the posterior half of the body, and (c) fertilized eggs collected within a few hours of their release into the brood pouch, has clearly demonstrated that before the fertilization the chorion is the only envelope present in the egg of oniscidean isopods. In the mature eggs, the chorion appears as a uniformly electron‐dense lamina, about 0.4–0.5 µm thick, which does not show any specialized area. A second envelope, described by other authors as vitelline envelope, is formed above the oolemma only right after fertilization and appears separated from the chorion by a space full of liquid. The ways in which the genesis of this envelope is realized are not yet clear; it could be interpreted rather as a fertilization membrane. The investigations carried out with the aid of a battery of FITC‐lectins have highlighted the presence at the chorion surface of unfertilized eggs of various saccharide residues distributed in uniform way. No significant change was observed in the pattern of lectins binding to the chorion of eggs taken from the brood pouch, thus demonstrating how, after the fertilization, no significant rearrangement in the distribution of saccharide residues present on the egg surface occurs in A. vulgare. The ways in which, therefore, the recognition, the binding and the entry of the peculiar sperm of oniscidean isopods into the egg occur, still remain all to be deciphered. Microsc. Res. Tech. 79:792–798, 2016.