Electrochemical Sensors for Direct Reagentless Measurement of Superoxide Production by Human Neutrophils

Abstract

Electrochemical sensors based on immobilised cytochrome c or superoxide dismutase for the measurement of superoxide radical production by stimulated neutrophils are described. Cytochrome c was immobilised covalently at a surface-modified gold electrode and by passive adsorption to novel platinised activated carbon electrodes (PACE). The reoxidation of cytochrome c at the electrode surface upon reduction by superoxide was monitored using both xanthine/xanthine oxidase and stimulated neutrophils as sources of the free radical. In addition, bovine Cu/Zn superoxide dismutase was immobilised to PACE by passive adsorption and superoxide, generated by xanthine/xanthine oxidase, detected by oxidation of hydrogen peroxide produced by the enzymic dismutation of the superoxide radical. A biopsy needle probe electrode based on cytochrome c immobilised at PACE and suitable for continuous monitoring of free radical production was constructed and characterised.

Simultaneous use of electrochemistry and chemiluminescence to detect reactive oxygen species produced by human neutrophils

“…On the other hand, addition of catalase increased the signal (Fig. 4D between the rate of the current increase from O 2 •-sensitive gold electrodes and the number of cells seeded was obtained (Chang et al, 2005a(Chang et al, , 2005b, whereas in the present work, and some other studies (McNeil et al, 1992(McNeil et al, , 1995, the response curve was not linear, but showed a saturation behaviour at higher neutrophil loadings, reflecting the attachment of the latter to the electrode surface.…”

Selective and sensitive electrochemical biosensing of superoxide anion production by biological systems : a short overview of recent trends

“…The most developed device is constituted of cytochrome ccoated gold [17][18][19][20][21][22][23][24][25][26][27] or platinized carbon electrode [21,28,29]. Superoxide anion reduces the immobilized ferricytochrome c, which is immediately re-oxidized by the surface-modified electrode at a potential of 15-25 mV vs Ag/AgCl, as illustrated in figure 1.…”

“…Another approach for making cytochrome c-coated electrodes consistes to achieve the immobilisation of cytochrome c by passive adsorption on platinized carbon electrode, PACE, as follow: a platinized activated carbon electrode (1 cm 2 area) is immersed in a phosphate buffer solution (pH = 7.4) of cytochrome c (10 mg/ml) and shaked gently for few hours [21]. After this time, the PACE incorporating adsorbed cytochrome c is rinsed throughly and stored in phosphate buffer solution at 5°C.…”

“…The sensitivity of these both biosensors to the superoxide anion was generally assessed by using the xanthine (Xa) / xanthine oxidase (XOD) reaction: .µM -1 [21]. The increase in sensitivity for the PACE configuration is thought to arise from the much larger effective surface area of PACE compared with the planar gold electrode.…”

In vivo Detection of Superoxide Anion Production by the Brain Using a Cytochrome c Electrode

“…Calibration of electrodes using xanthine-xanthine oxidase system in vitro Cytochrome c electrodes were prepared using a modi fication of the method of McNeil et al (1992). Electrodes were constructed of platinized carbon electrode (PACE) material (EFCG S type material; E-Tek, Natick, MA, U.S.A.), which was the generous gift of Dr. Calum Mc Neil.…”