Photorelease of caged Ca(2+) is a uniquely powerful tool to study the dynamics of Ca(2+)-triggered exocytosis from individual cells. Using photolithography and other microfabrication techniques, we have developed transparent microchip devices to enable photorelease of caged Ca(2+), together with electrochemical detection of quantal catecholamine secretion from individual cells or cell arrays as a step towards developing high-throughput experimental devices. A 100 nm thick transparent indium-tin-oxide (ITO) film was sputter-deposited onto glass coverslips, which were then patterned into 24 cell-sized working electrodes (approximately 20 microm by 20 microm). We loaded bovine chromaffin cells with acetoxymethyl (AM) ester derivatives of the Ca(2+) cage NP-EGTA and Ca(2+) indicator dye fura-4F, then transferred these cells onto the working ITO electrodes for amperometric recordings. Upon flash photorelease of caged Ca(2+), a uniform rise of [Ca(2+)](i) within the target cell leads to quantal release of oxidizable catecholamines measured amperometrically by the underlying ITO electrode. We observed a burst of amperometric spikes upon rapid elevation of [Ca(2+)](i) and a "priming" effect of sub-stimulatory [Ca(2+)](i) on the response of cells to subsequent [Ca(2+)](i) elevation, similar to previous reports using different techniques. We conclude that UV photolysis of caged Ca(2+) is a suitable stimulation technique for higher-throughput studies of Ca(2+)-dependent exocytosis on transparent electrochemical microelectrode arrays.

Here we describe a method to fabricate a multi-channel high-throughput microchip device for measurement of quantal transmitter release from individual cells. Instead of bringing carbon-fiber electrodes to cells, the device uses a surface chemistry approach to bring cells to an array of electrochemical microelectrodes. The microelectrodes are small and “cytophilic” in order to promote adhesion of a single cell whereas all other areas of the chip are covered with a thin “cytophobic” film to block cell attachement and facilitate movement of cells to electrodes. This cytophobic film also insulates unused areas of the conductive film, thus the alignment of cell docking sites to working electrodes is automatic. Amperometric spikes resulting from single-granule fusion events were recorded on the device and had amplitudes and kinetics similar to those measured using carbon-fiber microelectrodes. Use of this device will increase the pace of basic neuroscience research and may also find applications in drug discovery or validation.

Electrochemical measurement of transmitter or hormone release from individual cells on microchips has applications both in basic science and drug screening. High-resolution measurement of quantal exocytosis requires the working electrode to be small (cell-sized) and located in immediate proximity to the cell. We examined the ability of candidate electrode materials to promote the attachment of two hormone-secreting cell types as a mechanism for targeting cells for to recording electrodes with high precision. We found that nitrogen-doped diamond-like carbon (DLC:N) promoted cell attachment relative to other materials tested in the rank order of DLC:N > In2O3/SnO2 (ITO), Pt > Au. In addition, we found that treating candidate electrode materials with polylysine did not increase attachment of chromaffin cells to DLC:N, but promoted cell attachment to the other tested materials. We found that hormone-secreting cells did not attach readily to Teflon AF as a potential insulating material, and demonstrated that patterning of Teflon AF leads to selective cell targeting to DLC:N “docking sites”. These results will guide the design of the next generation of biochips for automated and high-throughput measurement of quantal exocytosis.

Abstract-The charge-storage characteristics of a metal-oxidesemiconductor (MOS) structure containing size-tunable sub-2 nm Pt nanoparticles (NPs) between Al 2 O 3 tunneling and capping oxide layers were studied. Significantly different amounts of memory window were obtained with the different sizes of Pt NP embedded MOS structures and reached a maximum of 4.3 V using a 1.14 nm Pt NP, which has the strongest charging capability caused by optimum size and the largest particle density obtained in our deposition method. Satisfactory long-term nonvolatility was attained in a low electric field due to the Coulomb blockade and quantum confinement effects in ∼1 nm Pt NP. These properties are very promising in view of device application.Index Terms-Nanoparticle (NP), nonvolatile memory (NVM), size-tunable platinum.

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