The Human Topoisomerase I Drug Screening Kit is a specialized assay system designed for the detection and quantification of compounds that target topoisomerase I (Top1) by inducing DNA nicks. This kit enables researchers to evaluate potential inhibitors that interfere with Top1 activity, particularly those that stabilize the cleaved intermediate complex.

The detection mechanism relies on trapping the Top1-DNA complex in its nicked intermediate state using protein denaturants, such as SDS. This process results in the formation of covalent Top1-DNA complexes, which contain nicked open circular DNA—a hallmark of Top1 inhibition. The presence of these nicked DNA intermediates can be visualized via Ethidium Bromide agarose gel electrophoresis.  These EB gels are ideal for detecting nicked plasmid DNA as described in the Kit protocol

Since trapping nicked intermediates occurs at low efficiency, the assay is optimized to detect the effects of Top1 poisons (aka IFP or interfacial Poisons) such as camptothecin, a natural product known to stabilize the Top1-DNA intermediate. This stabilization leads to an increase in nicked DNA products, forming the foundation of the mechanistic drug screening assay.

This drug screening kit is designed to allow the investigator to quickly identify novel inhibitors of Human Topoisomerase I (included with kits denoted -A in catalog numbers).