The isolation, purification and evaluation of the first thermal stable alkaline phosphatase to be isolated and purified: its evaluation in DNA based diagnostic tests. DNA based diagnostics are currently limited by the availability of enzyme based signal generation systems. Those available are from mesophillic sources. As a consequence these impose , upon the developed diagnostic tests, and absolute requirement for incorporation of the signal generation component at the end of all the manipulations. This proposal is designed to make available a new generation of signal generation molecules, that will allow incorporation during the procedure thus allowing multiple incorporation of the signal generation molecule. It is proposed that we make available thermostable enzymes, which will enhance both the signal produced as a result of the consequential enhanced kinetics of anu reaction and also allow the incorporation fo the enzymes during the diagnsotic test. DNA based diagnsotic tests reauire that steps are performed at elevated teperatures to allow denaturation and hybridisation to occur. Thermostable enzymes will be resstant ot these steps. In addition to these advantages the currently available enzymes have a limited purity activity and in the case of animal derived alkaline phosphatase, contain DNA and RNA ases which destroy the probe molecules.
Funding SchemeEAW - Exploratory awards