Raman micro-spectroscopy combined with multivariate analysis was employed to monitor real-time biochemical changes induced in living cells in vitro following exposure to a pharmaceutical. The cancer drug etoposide (topoisomerase II inhibitor) was used to induce double-strand DNA breaks in human type II pneumocyte-like cells (A549 cell-line). Raman spectra of A549 cells exposed to 100 microM etoposide were collected and classical least squares (CLS) analysis used to determine the relative concentrations of the main cellular components. It was found that the concentrations of DNA and RNA significantly (P < 0.05) decreased, whilst the concentration of lipids significantly (P < 0.05) increased with increasing etoposide exposure time as compared to control untreated A549 cells. The concentration of DNA decreased by 27.5 and 87.0% after 24 and 48 h exposure to etoposide respectively. Principal components analysis (PCA) successfully discriminated between treated and untreated cells, with the main variance between treatment groups attributed to changes in DNA and lipid. DNA fragmentation was confirmed by Western blot analysis of apoptosis regulator protein p53 and cell metabolic activity determined by MTT assay. The over-expression of p53 protein in the etoposide treated cells indicated a significant level of DNA fragmentation and apoptosis. MTT tests confirmed that cellular metabolic activity decreased following exposure to etoposide by 29.4 and 61.2% after 24 and 48 h, respectively. Raman micro-spectroscopy may find applications in the toxicology screening of other drugs, chemicals and new biomaterials, with a range of cell types.

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The initial adsorption of proteins to materials for implantable microelectrodes is an important factor in determining the longevity and stability of the implant. Implantable microelectrodes have the potential to become part of neural prosthesis to restore lost nerve function after nerve damage. The aim of this study was to identify an optimum material for electrode recording sites on implantable microelectrodes. Common materials for electrode sites are gold, platinum, iridium, and indium tin oxide; these, along with a reference material (titanium) were investigated. Once an implant is in the body, protein adsorption takes place almost instantly before the cells reach the surface of an implant. In this project, the adsorption of proteins to materials for electrode sites has been investigated using atomic force microscopy and ellipsometry.The protein layers adsorbed to Au after 7 and 28 days were the smoothest and the thinnest compared to all the other substrate materials. Among all the materials studied, Au is the best conductor, the most readily available and has good adhesion strength. Au is the material of choice for electrode recording sites on implantable microelectrodes. This work has demonstrated the suitability of different materials for electrode sites on implantable microelectrodes.

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