High-speed scanning electrochemical microscopy method for substrate kinetic determination: method and theory.

Scanning electrochemical microscopy (SECM) allows imaging and analysis of a variety of biological samples, such as living cells. Up to now, it still remains a challenge to successfully decouple signals related to topography and reactivity. Furthermore, such delicate samples require careful adjustment of experimental parameters, such as scan velocity. The present study proposes a method to extract a substrate's kinetic rate by numerical modeling and experimental high speed constant height SECM imaging. This is especially useful for the determination of substrates with unknown surface reaction kinetics and large topographical features. To make this approach applicable to soft cell samples, which cannot be imaged at high velocity, a nonlinear fit strategy is presented to obtain kinetic rate values also under slow scan velocity conditions.

Fabrication of carbon, gold, platinum, silver, and mercury ultramicroelectrodes with controlled geometry.

A simple, fast, and reproducible method for the fabrication of disk ultramicroelectrodes (UMEs) with controlled geometry is reported. The use of prepulled soda-lime glass capillaries allows one to bypass the irreproducible torch-sealing and experimentally challenging tip-sharpening steps used in conventional fabrication protocols. A micron-sized electroactive wire is sealed inside this capillary producing UMEs with a highly reproducible geometry. Total fabrication time (1 h) and experimental difficulty are significantly reduced. Disk UMEs with various diameters and cores were fabricated, including carbon fiber (7 and 11 µm), gold (10 and 25 µm), platinum (10 and 25 µm), silver (25 µm), and mercury (25 µm). The ratio of the insulating sheath to the electroactive core of the UMEs was 2.5-3.6. Silver UMEs were also used to produce a Ag/AgCl microreference electrode. This general fabrication method can readily be applied to other electroactive cores and could allow any research group to produce high quality disk UMEs, which are a prerequisite for quantitative scanning electrochemical microscopy.

Disk-shaped amperometric enzymatic biosensor for in vivo detection of D-serine.

At the synapse, D-serine is an endogenous co-agonist for the N-methyl-D-aspartate receptor (NMDAR). It plays an important role in synaptic transmission and plasticity and has also been linked to several pathological diseases such as schizophrenia and Huntington's. The quantification of local changes in D-serine concentration is essential to further understanding these processes. We report herein the development of a disk-shaped amperometric enzymatic biosensor for detection of D-serine based on a 25 µm diameter platinum disk microelectrode with an electrodeposited poly-m-phenylenediamine (PPD) layer and an R. gracilis D-amino acid oxidase (RgDAAO) layer. The disk-shaped D-serine biosensor is 1-5 orders of magnitude smaller than previously reported probes and exhibits a sensitivity of 276 µA cm(-2) mM(-1) with an in vitro detection limit of 0.6 µM. We demonstrate its usefulness for in vivo applications by measuring the release of endogenous D-serine in the brain of Xenopus laevis tadpoles.