Enzymes and Cells Confined in Silica Nanopores
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Enzymes and Cells Confined in Silica Nanopores
Jacques Livage, Cécile Roux, Thibaud Coradin, Souad Fennouh, Stéphanie Guyon, Laurie Bergogné, Anne Coiffier and Odile Bouvet1 Laboratoire de Chimie de la Matière Condensée, Université Pierre et Marie Curie, 4 place Jussieu, 75252 Paris, France 1 Unité des Entérobactéries, Institut Pasteur, 28 rue du Dr Roux, 75724 Paris, France
ABSTRACT The sol-gel process opens new possibilities in the field of biotechnologies. Sol-gel glasses are formed at room temperature via the polymerization of molecular precursors. Enzymes can be added to the solution of precursors and trapped within the growing silica network. Small substrate molecules can diffuse through the pores allowing reactions to be performed in-situ, within the silica gels. Enzyme are encased by the hydrated silica in a cage tailored to their size, they retain their biocatalytic activity and may even be stabilized within the sol-gel matrix. Whole cell bacteria have also been immobilized within sol-gel glasses. They behave as a "bag of enzymes" and their membrane protects enzymes against denaturation and leaching. The cellular organization of bacteria cells is preserved upon encapsulation. Experiments performed with Escherichia coli induced to β-galactosidase show that they still exhibit noticeable enzymatic activity. Some degradation of the cell walls may even occur increasing the "measured" activity. However silica gels made from aqueous precursors seem to prevent bacteria from natural degradation upon ageing. Antibody-antigen recognition has been shown to be feasible within sol-gel matrices. Trapped antibodies bind specifically the corresponding haptens and can be used for the detection of traces of chemicals. Even whole cell protozoa have been encapsulated without any alteration of their cellular organization. For medical applications, trapped parasitic protozoa have been used as antigens for blood tests with human sera. Antigen-antibody interactions were followed by the so-called Enzyme Linked ImmunoSorbent Assays (ELISA).
INTRODUCTION Entrapment in crosslinked organic polymers is a well known method for the immobilization of enzymes and whole cells. Entrapped biomolecules are physically confined within the polymer matrix and can be reused several times. Organic polymers such as polyacrylamide gels are currently used in biotechnology but silica glasses could offer some advantages such as improved mechanical strength and chemical stability. Moreover they don't swell in aqueous or organic solvents preventing leaching of entrapped biomolecules. However glasses are made at high temperature and, up to now, enzyme immobilization can only be performed via adsorption or covalent binding onto the surface of porous glasses [1]. The so-called sol-gel process opens new possibilities in the field of biotechnology [2-4]. Solgel glasses are formed at room temperature via the polymerization of molecular precursors such T1.4.1
as metal alkoxides. Proteins can be added to the solution of precursors. Hydrolysis and condensation t
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