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N. V. Komarova, M. S. Andrianova, M. I. Saveliev, A. E. Kuznetsov
Optimization
of silicon dioxide surface functionalization protocol for design of receptor
layer of the biosensor for detection of explosives
Abstract
The development of the receptor layer of the
biosensor for detection of explosive compounds is described. Covalent
modification was chosen for immobilization of E. coli nitroreductase on
the gate oxide of ion-sensitive field effect transistor (ISFET) that was
comprised of silicon dioxide. Self-assembled monolayer technique was used for
immobilization. This method assumes the usage of different silanes and spacer
molecules for activation of SiO2 surface. Two different
immobilization strategies were compared, one using asymmetric spacers
(3-maleimidobenzoic acid N-hydroxysuccinimide ester and 4-(4-maleimidophenyl)butyric
acid N-hydroxysuccinimide ester) and another using symmetric glutaric
dialdehyde linker accompanied with appropriate silanes. For the first method,
the dependence of functionalization efficiency on silane concentration was
studied. The sufficient density of enzyme molecules on the surface of SiO2
was reached at 0.0015% of silane concentration. The type of asymmetric linker
had no influence on immobilization efficiency. The method implying glutaric
dialdehyde resulted in higher activity of the immobilized enzyme. For this
method, immobilization procedure was optimized. The method was adopted for
immobilization on E. coli nitroreductase inside the channel of
microfluidic system above ISFET surface. For this purpose
(3-aminopropyl)triethoxysilane was changed to the corresponding silatrane and
enzyme concentration was increased to 30 µg/ml. The optimized procedure was
successfully used for the development of the biosensor for explosives
detection.
Key words: enzyme,
immobilization, self-assembled monolayers, biosensor.
Copyright (C) Chemistry Dept., Moscow State University, 2002
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