The invention includes methods of detecting glycosylases. A test sample is mixed with substrate polynucleotide. A primer and a polymerase are added. An endonuclease is provided and a probe oligonucleotide sequence labeled with first and second labels is utilized for detection. The invention includes N-glycosylase assay methods. A test sample is mixed with substrate polynucleotide and formation of an abasic site is detected by forming a product that is complementary to a portion of the substrate sequence ending at the abasic site. The product is dissociated and is extended utilizing a polymerase. A probe is hybridized to the product and is cleaved. The invention includes synthetic substrates, transcription primers and probe molecules. The invention also includes an N-glycosylase detection kit including a substrate polynucleotide, an endonuclease and a dual-labeled probe having a fluorescent label and a quencher moiety.
 The United States Government has certain rights in this invention pursuant to Contract No. DE-AC07-05ID14517 between the United States Department of Energy and Battelle Energy Alliance, LLC.