Abstract
To develop an in situ detection system of 2-acetylaminofluorene(AAF)-DNA adducts, we tried to establish monoclonal antibodies against them. Mice were immunized with AAF-ssDNA coupled with protein, and then the spleen cells were fused with myeloma cells. Out of 359 hybridoma cells, 5 produced antibodies showing preferential binding to AAF-DNA than to DNA. After clonings, 5 types of monoclonal antibodies were obtained. The antibodies showed the high binding to AAF-DNA, but undetectable or minimal binding to undamaged DNA or UV-irradiated DNA. The competitive inhibition experiments revealed that the epitope is dG-C8-AAF in DNA, but deacetylated dG-C8-AF is also recognized with less efficiency. Using the most promising antibody AAF-1, we could measure the formation of AAF-DNA adducts in DNA of xeroderma pigmentosum group A (XP-A) cells following exposure of N-acetoxy-2-AAF (25-150 uM). Moreover, we could detect AAF-DNA adducts in XP-A cells in situ. Thus, we succeeded in establishing, for the first time, the monoclonal antibodies applicable to in situ detection of AAF-DNA adducts. [J Radiat Res 44:440 (2003)]
