Immobilization strategies/techniques

Physical or electrochemical adsorption uses non-covalent forces to affix the nucleic acid to the solid support and represents a relatively simple mechanism for attachment that is easy to automate. Adsorption was favoured and described in some chapters as suitable immobilization technique when multisite attachment of DNA is needed to exploit the intrinsic DNA oxidation signal in hybridization reactions. Dendrimers such as polyamidoamine with a high density of terminal amino groups have been reported to increase the surface coverage of physically adsorbed DNA to the surface. Furthermore, electrochemical adsorption is described as a useful immobilization strategy for electrochemical genosensor fabrication. 

As detailed in this overview, the non-covalent attachment of catalysts on a solid support is an important additional technique for the separation and recovery of catalysts from reaction mixtures. Such non-covalent immobilization strategies bring together a number of advantages of solution-phase chemistry and solid-phase supported chemistry. The catalysts can be separated from reaction mixtures by simple filtration. The pre-catalysts can be prepared and characterized in solution. The underlying principle is partitioning between a solid phase or a supported liquid phase and a liquid reaction phase of different solvating power. 

Dehydrogenases are usually multimeric enzymes. Thus, under certain conditions even thermophilic dehydrogenases may be easily inactivated by subunit dissociation, being these enzymes an excellent target for the development of stabilization strategies by immobilization and post-immobilization modification techniques as shown in Fig. 6.4.5. 

The degree of enzyme purity will ultimately affect fuel cell performance, particularly when enzyme preparations are used to form immobilized films on electrode surfaces in DET reactions. Contaminating proteins that do not provide electron transfer effectively foul the electrode. When enzyme immobilization techniques are specific to the enzyme, then enzyme purity may not be as much as an issue, but rarely the immobilization technique is absolutely specific to the cathodic or anodic enzyme. For example, an attractive immobilization strategy is to link a particular enzyme to an electrode via its cofactor (e.g., flavin adenine dinucleotide (FAD), nicotinamide adenine dinucleotide (NAD), etc.) [59]. The cofactor is linked to the electrode material first and then the apoenzyme is allowed to naturally bind to the cofactor all other proteins in the enzyme preparation that cannot bind the cofactor remain unbound and can be removed. Enzymes used in fuel cells are not so unique, and proteins in the immobilizing preparation may use the same cofactor but not the same fuel during fuel cell analysis or operation. 

A variety of approaches exist for stabilizing proteins, for example, chemical modification, immobilization, and site-directed mutagenesis [95,96], but these techniques are not within the scope of this chapter. The focus here will be on stabilization of proteins via formulation development. The principal formulation strategy is to stabilize the protein using clinically acceptable additives (excipients) or through the use of suitable pharmaceutical-processing technologies. 

The above example outlines a general problem in immobilized molecular catalysts - multiple types of sites are often produced. To this end, we are developing techniques to prepare well-defined immobilized organometallic catalysts on silica supports with isolated catalytic sites (7). Our new strategy is demonstrated by creation of isolated titanium complexes on a mesoporous silica support. These new materials are characterized in detail and their catalytic properties in test reactions (polymerization of ethylene) indicate improved catalytic performance over supported catalysts prepared via conventional means (8). The generality of this catalyst design approach is discussed and additional immobilized metal complex catalysts are considered. 

Immobilization techniques have been applied in the preparation of immobilized CL reagents, with specific advantages such as reusability, improved stability, and increased efficiency. These strategies have been applied in the development of CL sensors, which today constitute the most important tools in analytical chemistry because of the high sensitivity offered. Optical fibers have been used to transfer light in order to improve the quality of detection, and new types of flow-through cells have been introduced in the construction of CL sensors. Also, selectivity has been considerably improved by the utilization of enzymatic or antigen-antibody reactions. 

Conventional pump-and-treat techniques are not very effective in restoring aquifers impacted by DNAPLs. This ineffectiveness is a result of the relatively low solubility of the DNAPL and the large capillary forces that immobilize the nonaqueous phase. Over the past decade, several innovative and experimental strategies have been tested for more effective recovery of DNAPLs. These strategies include the more conventional use of surfactants, and thermally enhanced extraction or steam injection. Other more experimental approaches include cosolvent flooding and density manipulations. Each of these approaches is discussed below. 

Polyoxometalates undoubtedly have enormous catalytic potential in liquid phase selective oxidation of organic compounds. Various strategies for immobilization of POMs on solid matrices have been developed during the past two decades and opened new opportunities for practical applications. The most developed and widely used technique is electrostatic... 

Flowever, the focus of the major part of the chapters lies on the couphng chemistry used for DNA immobilization. Successful immobihzation techniques for DNA appear to either involve a multi-site attachment of DNA (preferentially by electrochemical and/or physical adsorption) or a single-point attachment of DNA (mainly by surface activation and covalent immobihzation or (strept)avidin-biotin linkage). Immobilization methods described here comprise physical or electrochemical adsorption, cross-linking or entrapment in polymeric films, (strept)avidin-biotin complexation, a surface activation via self-assembled monolayers using thiol linker chemistry or silanization procedures, and finally covalent coupling strategies. 

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