Venera Aiello

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.

Immobilization of the Enzyme Glucose Oxidase on Both Bulk and Porous SiO2 Surfaces

Silicon dioxide surfaces, both bulk and porous, were used to anchor the enzyme glucose oxidase. The immobilization protocol was optimized and the samples characterized using X-ray Photoelectron Spectroscopy, Energy Dispersive X-rays coupled to scanning electron microscopy and enzymatic activity measurements. We show that a uniform layer was obtained by activating the oxide before immobilization. X-ray Photoelectron Spectroscopy measurements carried out on bulk oxide showed that the silicon substrate signal was fully screened after the enzyme deposition showing the absence of uncovered surface regions. The enzyme presence was detected monitoring both the C 1s and N 1s signals. Finally, enzymatic activity measurements confirmed that the glucose oxidase activity was preserved after immobilization and maintained after three months of shelf life if the sample was properly stored. The importance of using porous silicon oxide to maximize the surface area was also evidenced.

On the Relationship between Jetted Inks and Printed Biopatterns: Molecular-Thin Functional Microarrays of Glucose Oxidase

Arrays of circular spots of glucose oxidase have been obtained on functionalized silicon oxide by piezoelectric inkjet printing and the enzymatic activity toward glucose recognition has been monitored. The addition of glycerol to the molecular ink allows to obtain high spot definition and resolution (tens of micrometers wide; one molecule tall), but in spite of its well-known structural stabilizing properties, in dynamic conditions it may lead to increased protein stresses. The jetting voltage and pulse length have been found to be critical factors for both activity retention and pattern definition. High voltages and pulse lengths results in stress effects along with the loss of activity, which, at least in our experimental conditions, has been found to be recovered in time.

From Atomistic to Device Level Investigation of Hybrid Redox Molecular/Silicon Field-Effect Memory Devices

In this paper, an extensive investigation of hybrid molecular/Si field-effect memories is presented, where redox ferrocene (Fc) molecules play the role of the memory charge storage nodes. Engineering of the organic linkers between Fc and Si is achieved by grafting Fc with different linker lengths. The study shows a clear correlation between results from atomistic computational density functional theory, electrochemical measurements (cyclic voltammetry) and electrical data obtained by a detailed study on capacitors and pseudo-MOS devices. Physical-chemical analyses (atomic force microscopy, high-resolution transmission electron microscopy, and X-ray photoelectron spectroscopy), corroborate the quality of molecular layers on devices.

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.