This paper describes laser-based methods for preparing micropatterns of bioactive molecular species in self-assembled monolayers (SAMs) and micropatterns of proteins and other biological molecules immobilized on solid substrates. Applications of these micropatterned surfaces in multianalyte biosensing and tissue engineering are emphasized. The focus of the paper is on the use of a computer-controlled laser ablation system comprising a research-grade inverted optical microscope, a pulsed nitrogen-pumped dye laser emitting at 390 nm, a programmable sample stage, and the computerized control system. The laser system can be implemented in a typical biosensor or tissue culture laboratory to enable the facile and reproducible fabrication of micropatterned surfaces by several methods. Various methods for patterning are discussed with examples given and emphasis placed on (1) laser ablation in the fabrication of photolithography masks, (2) electrochemical patterning of SAMs, and (3) laser desorption of SAMs. The relative merits of each technique are discussed with respect to application in fabrication of active surfaces for biosensing and tissue culture applications.
A new enzyme electrode concept featuring glucose oxidase (GOD) immobilized on fine carbon powder in a fluid state and a cross-linked GOD enzyme layer has been developed. This enzyme electrode has been tested in vitro at 37°C and has a lifetime of 3 months after which it can be recharged with fresh enzyme. In vitro interference tests carried out on this sensor used with cellulose acetate membranes are described. Sulfur-free EDTA-treated cellulose acetate membranes [molecular weight cutoff at 1000) were used, some of which were coated with positively and negatively charged hydrogel layers. The sensors showed a stable and linear response to glucose concentrations 5300 mg/ dL, in the presence of glucose alone in the phosphate buffer medium and in the presence of interferences. The effects of ascorbic acid, bilirubin. creatinine, L-qstine, glycine. uric acid, and urea on the amperometric signal of the sensor were studied. The cellulose acetate membrane coated with the negatively charged hydrogel layer provided good protection for the e n q m e electrode, especially in the presence of uric acid. bilirubin, and L-qstine.
Self-assembled monolayers (SAMs) of alkylsiloxanes were formed from hexyltrichlorosilane (HTS) and octadecyltrichlorosilane (OTS) on surfaces of thin films of a complex oxide, lead zirconate titanate (PZT). X-ray photoelectron spectroscopy (XPS) and contact angle measurements confirmed the formation of a thin, uniform organic layer on the surface of the PZT, consistent with the hypothesis that a densely packed organic monolayer is formed on the PZT. Angle-resolved high-resolution XPS suggested that the surface of the PZT thin film includes a top layer deficient in titanium and consisting mainly of oxides of lead and zirconium, along with hydroxylated zirconium that may react with alkyltrichlorosilanes to form the SAMs. The effect on these SAMs of exposure to acidic media was probed by wettability measurements, XPS, and scanning electron microscopy (SEM). Contact angle measurements with water and hexadecane indicated that the SAMs formed from the longer alkylsilane, OTS, were stable in HCl over long periods of time (at least 3 days), while the SAMs formed from the short-chain alkylsilane, HTS, degraded after 12 h. The XPS spectra of SAMs formed from OTS and exposed to HCl solution were similar to those obtained for similar SAMs not exposed to HCl. SEM also confirmed that the SAMs formed from OTS can act as protective barriers for PZT against etching by HCl.
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