During apoptosis, DNA is fragmented following the apoptotic activation of intracellular endonucleases. The In Situ Apoptosis Detection Set is designed to detect fragmented DNA histochemically by TUNEL (TdT-mediated dUTP Nick End Labeling). Via this method, fluorescein-labeled nucleotides are incorporated in situ onto the 3' ends of DNA fragments, allowing histologic localization and detection of individual apoptotic cells.
Apoptosis plays a significant role in various stages of cancer as well as cell development, growth and proliferation. Physical (e.g., radiation, heat) and chemical (e.g., medicine) factors can also induce the apoptotic state. One of the features of apoptotic cells is the cleavage of nuclear DNA to approximately 185 bp fragments. This fragmented DNA can be detected histochemically by the TUNEL method.
Principle
TUNEL, an acronym for terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling, is an effective method for measuring DNA fragments resulting from the apoptotic activation of intracellular endonucleases. The TdT enzyme is used to label the 3'-OH ends of DNA fragments with fluorescein-dUTP. These fluorescence-labeled nucleotides are incorporated in situ onto DNA fragment ends, allowing histologic localization of individual cells. As a result, cells undergoing apoptosis can be easily detected by fluorescence microscopy. Since incorporated fluorescein-dUTP can also be detected using peroxidase-labeled anti-fluorescein antibody, it is possible to detect apoptotic cells using light microscopy.
