Enzymes

Owing to their ability to catalyze specific reactions yielding distinct, detectable products, enzymes are probably the most popular molecular tag for expression of recombinant genes. The most commonly used enzymes include, chloramphenicol acetyltransferase (CAT), firefly luciferase; P-galactosidase, alkaline phosphatase; and (3-glucuronidase Some of the key reasons for selecting these enzymes as functional labels include high signal-to-background ratios of the catalyzed reactions, high stability of enzyme activity, and the high sensitivity for detection A number of methods are currently in use for the detection of enzyme activity, including standard colorimetric assays, more sensitive fluorescence- or lummescence-based procedures, chromogenic histochemistry, and immunohistochemistry or solution-phase immunoassays such as radioimmunoassay or enzyme-linked immunosorbent assay (ELISA). The specific characteristics of some of these enzymes and their respective detection protocols are presented m detail m a number of chapters in this book (Chapters 3, 4, 5, and 6).

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