CORE--ANTIOXIDANT ENZYME

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DESCRIPTION (provided by applicant): The purpose of the Antioxidant Enzyme Core (AEC) is to provide antioxidant enzyme expertise and analysis for the individual research projects. Discussion between a project investigator and the AEC Director will be held to determine which experiments and/or assays are run. The AEC staff will carry out the proposed work or will advise the investigator on how to perform the experiments. Some of this work, particularly transfections of new cDNAs or in new cell lines as well as the development of antisense reagents, will be developmental in nature. The AEC will aid in enzyme activity assays, western, northern, and Southern analysis. The enzyme assays will be run on the antioxidant enzymes copper-and zinc-containing superoxide dismutase (CuZnSOD), manganese-containing superoxide dismutase (MnSOD), catalase (CAT), and glutathione peroxidase (GPx, both cytosolic and phospholipid). For Project 1, work will also be done on the white cell enzyme myeloperoxidase (MPO). For reagents, we have antibodies and cDNA probes to all primary antioxidant enzymes. In addition, the AEC will aid in transfecting the cDNAs for these enzymes into cell lines of interest to the project investigators. We have experience with several methods of transfection, as well as many different expression vectors for these enzymes. In the course of this work, the AEC will continue to maintain and develop reagents i.e., antibodies, cDNAs, and expression vectors for the antioxidant proteins. The AEC will also have as a goal the development of antisense reagents to inhibit the levels of all of the major antioxidant proteins.