The objective of this project is to utilize currently available molecular biological techniques to develop a sensitive and practical test of exposure to genotoxic agents. A set of enzymatic treatments of the human alphoid DNA sequence will be tested in the laboratory for efficacy in detecting abnormalities induced by irradiating T-lymphocytes in freshly drawn blood. Primary emphasis will be placed on procedures which lead to the cleavage of aberrant DNA while leaving the normal 342 base pair alphoid sequence intact. Preliminary experiments will employ P-32 labeled nucleotides and autoradiography of electrophoresis gels to assess cleavage rates. As the procedure is optimized, less energetic labels will be employed and cleavage rates will be determined by liquid scintillation counting of two portions of the electrophoresis gel. The utility of High Pressure Liquid Chromatography will be investigated as an alternative for electrophoresis in routine applications. After the mechanics of the technique have been optimized, a limited number of observations will be made on (presumably) minimally exposed members of the general population in order to estimate the residual background damage rate as measured by the procedure. Additional measurements will be made on blood samples from individuals with documented elevated occupational or therapeutic radiation exposures. An analogous procedure for use in a suitable laboratory animal will be sought in order that in vivo animal experiments can be used to supplement in vitro exposure studies. The utility of the procedure for assessing exposures to a selected group of chemical genotoxins will then be evaluated.
