A novel approach is proposed for detection and analysis of apoptotic DNA damage in tissue sections, as well as in vivo. The technology is based on it situ ligation of DNA probes that will uniquely bind to characteristic DNA breaks, which other and we have demonstrated are a specific signature for apoptosis. Preliminary results using the first generation of the probes documented higher specificity for apoptosis detection compared with the currently used terminal transferase-based labeling (TUNEL), thus demonstrating the potential for development into a marketable kit for use in tissue sections. Probes under development include loopless hairpin oligoprobes with greater specificity and less background staining, as well as probes utilizing Fluorescence Resonance Energy Transfer (FRET) technology, which change color upon successful ligation. A much more powerful application of technology is related to in vivo imaging of apoptosis. In preliminary experiments we have tested the modification of the approach allowing it to detect specific DNA damage in vivo. Using this modified approach, we will develop a marketable in vivo detection procedure for analysis of apoptotic damage in brain, with visualization utilizing magnetic resonance imaging (MRI).
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