Interdigital 50 nm Ti Electrode Arrays Fabricated Using XeF2 Enhanced Focused Ion Beam Etching

The fabrication of interdigitated titanium nanoelectrode arrays of 50 nm in width and spacing is described in this work. The nanoarrays have been realized using a Ga+ focused ion beam (FIB). FIB milling is typically accompanied by redeposition of removed material, which represents an important hindrance for milling closely spaced nanostructures. Redeposition effects have been reduced by means of XeF2 gas assistance, which increases the etch yield by a factor of seven compared with pure ion milling. Furthermore, we used a simple adsorption model, which led to the conclusion that dwell time and refresh time should be 30 ms, respectively, for optimized XeF2 assisted Ti milling. The measured resistance R of the electrodes is higher than 1 G ohm

Ultrasensititve system for the real time detection of H2O2 based on strong coupling in a bio-plasmonic system

We theoretically investigate the dependence of the different parameters of an optical biosensor for the detection of Hydrogen peroxide (H2O2) based on absorption enhancement of Cytochrome c molecules near gold nanoparticles. H2O2 is a major reactive oxygen species which is involved in signaling pathways and oxidative stress in cells. We use the Green's function approach as well as confirm the corresponding simulation results using the surface integral formulation. Further we show that this technique can be applied for detection of other small molecules, like oxygen and carbon monoxide

Biophotonic tool for sensing the dynamics of H2O2 extracellular release in stressed cells

Hydrogen peroxide (H2O2) is known to play a multifaceted role in cell physiology mechanisms involving oxidative stress and intracellular signal transduction. Therefore, the development of analytical tools providing information on the dynamics of H2O2 generation remains of utmost importance to achieve further insight in the complex physiological processes of living cells and their response to environmental stress(1). In the present work we developed a novel optic biosensor that provides continuous real-time quantification of the dynamics of the hydrogen peroxide release from cells under oxidative stress conditions. The biosensor is based on the ultra-sensitive dark field optical detection of cytochrome c (cyt c) that exhibits a narrow absorption peaks in its reduced state (Fe(II)) at lambda = 550 nm. In the presence of H2O2 the ferrous heme group Fe(II) is oxidised into Fe(III) providing the spectroscopic information exploited in this approach. Extremely low limit-of-detection for H2O2 down to the subnanomolar range is achieved by combining scattering substrates (eg. polystyrene beads) able to shelter cyt c and an inverted microscope in dark field configuration. The developed biosensor was able to perform real-time detection of H2O (2) extracellular release from human promyelocytic leukemia cells (HL-60) exposed to lipopolysaccaride (LPS) that elicits strong immune-response. This biosensing tool is currently being implemented to the real-time detection of superoxide anion (O-2(center dot-)) and offers the possibility to extend to further oxidative stress biomarkers such as glutathione. More generally, multianalyte and dynamic informations might bring new insights to understand complex cellular metabolisms involved in oxidative-stress-related diseases and cytotoxic responses

Polarization sensitive silicon photodiodes using nanostructured metallic grids

In this paper, we present the design, fabrication, and characterization of wire grid polarizers. These polarizers show high extinction ratios and high transmission with structure dimensions that are compatible with current complementary metal-oxide-semiconductor (CMOS) technology. To design these wire grids, we first analyze the transmission properties of single apertures. From the understanding of a single aperture, we apply a modal expansion method to model wire grids. The most promising grids are fabricated on both a glass substrate and CMOS photodiode. An extinction ratio higher than 200 is measured

Gβγ and the C Terminus of SNAP-25 Are Necessary for Long-Term Depression of Transmitter Release

Short-term presynaptic inhibition mediated by G protein-coupled receptors involves a direct interaction between G proteins and the vesicle release machinery. Recent studies implicate the C terminus of the vesicle-associated protein SNAP-25 as a molecular binding target of Gβγ that transiently reduces vesicular release. However, it is not known whether SNAP-25 is a target for molecular modifications expressing long-term changes in transmitter release probability.This study utilized two-photon laser scanning microscopy for real-time imaging of action potential-evoked [Ca(2+)] increases, in single Schaffer collateral presynaptic release sites in in vitro hippocampal slices, plus simultaneous recording of Schaffer collateral-evoked synaptic potentials. We used electroporation to infuse small peptides through CA3 cell bodies into presynaptic Schaffer collateral terminals to selectively study the presynaptic effect of scavenging the G-protein Gβγ. We demonstrate here that the C terminus of SNAP-25 is necessary for expression of LTD, but not long-term potentiation (LTP), of synaptic strength. Using type A botulinum toxin (BoNT/A) to enzymatically cleave the 9 amino acid C-terminus of SNAP-25 eliminated the ability of low frequency synaptic stimulation to induce LTD, but not LTP, even if release probability was restored to pre-BoNT/A levels by elevating extracellular [Ca(2+)]. Presynaptic electroporation infusion of the 14-amino acid C-terminus of SNAP-25 (Ct-SNAP-25), to scavenge Gβγ, reduced both the transient presynaptic inhibition produced by the group II metabotropic glutamate receptor stimulation, and LTD. Furthermore, presynaptic infusion of mSIRK, a second, structurally distinct Gβγ scavenging peptide, also blocked the induction of LTD. While Gβγ binds directly to and inhibit voltage-dependent Ca(2+) channels, imaging of presynaptic [Ca(2+)] with Mg-Green revealed that low-frequency stimulation only transiently reduced presynaptic Ca(2+) influx, an effect not altered by infusion of Ct-SNAP-25.The C-terminus of SNAP-25, which links synaptotagmin I to the SNARE complex, is a binding target for Gβγ necessary for both transient transmitter-mediated presynaptic inhibition, and the induction of presynaptic LTD

FABRICATION OF NANOPORES IN SIN MEMBRANES USING FIB

Pressing adhering live cells onto the nanostructured SiN substrate and subsequent removal, planar cell membrane sheets (CMSs) were transferred in a well-defined orientation onto the SiN support. Using perforated thin SiN membranes both, extracellular and intracellular surfaces of CMSs become simultaneously accessible by targeting membrane constituents side-specifically with fluorescent markers. The perforation of the membranes has been performed using FIB technique

INTERDIGITATED NANOELECTRODES FABRICATED USING FIB

The use of interdigitated electrodes of nanometer scale establishes a broad field of new applications particularly in biotechnology. It allows to create electric field gradients at submicrometer scale. The goal of the project is the fabrication and characterization of nanoelectrodes in a sub 100 nm range using FIB-technology

Direct Anchoring of Cytochrome c onto Bare Gold Electrode for Sensing Oxidative Stress in Aquatic Cells

A novel third generation biosensor was developed based on one-shot adsorption of chemically-modified cytochrome c (cyt c) onto bare gold electrodes. The introduction of short-chain thiol derivatives (mercaptopropionic acid, MPA) on the lysine residues of cyt c enabled the very fast formation (<5 min) of an electroactive biological self-assembled monolayer (SAM) exhibiting a quasi-reversible electrochemical behavior and a fast direct electron transfer (ET). The heterogeneous ET rate constant was estimated to be ks = 1600 s-1, confirming that short anchors facilitate the ET via an efficient orientation of the heme pocket. In comparison, no ET was observed in the case of native and long-anchor (mercaptoundecanoic acid, MUA) modified cyt c directly adsorbed on gold. However, in both cases the ET was efficiently restored upon in-bulk generation of gold nanoparticles which acted as electron shuttles. This observation emphasizes that the lack of electroactivity might be caused by either an inappropriate orientation of the protein (native cyt c) or a critical distance (MUA-cyt c). Finally, the sensitivity of the bare gold electrode directly modified with MPA-cyt c to hydrogen peroxide (H2O2) was evaluated by amperometry and the so-made amperometric biosensor was able to perform real-time and non-invasive detection of endogeneous H2O2 released by unicellular aquatic microorganisms, Chlamydomonas reinhardtii, upon cadmium exposure

NANOSCALE DISPENSING OF LIQUIDS WITH AFM-CANTILEVER

The demand for tools suitable for deposition of small amount of material is continuously increasing. A standard SiN cantilever was modified in order to dispense single droplets of femto- and ato-liter (NAnoscale DISpensing (NADIS)). In order to open a nano fluid channel, holes down to 100 nm were milled in the apex of the cantilever using FIB. The liquid to dispense is loaded on the top-side of the cantilever extremity