Leírás
Termékismertető megtekintése (angol)
Apoptosis is a basic biological phenomenon of cells. It plays an important role in evolution, homeostasis and system development of organism. Some morphological, physiological or biochemical changes will happen during the apoptosis of cells, such as cell shrinkage, loss of contact between neighboring cells, loss of mitochondrial membrane potential, abnormal membrane permeability, chromatin condensation, nuclear fragmentation, DNA degradation; formation of membrane protrusions; phosphatidylserine’s turning out of membrane while the structure of cell membrane is complete. Finally, the cell breaks apart into multiple vesicles called apoptotic bodies, which will undergo phagocytosis. The above described morphological changes occur at different stages of the apoptosis.
A landmark of apoptosis is degradation of chromosome DNA. This specific and regular degradation produces DNA fragments in different lengths of an integer multiple of 180 bp to 200 bp. This is exactly the length of DNA strand that wraps histone. It suggests that the chromosomal DNA is cleaved at the junction between nucleosomes, producing oligonuclear fragments of different lengths. Experiments have shown that the regulated degradation of DNA is a result of an endogenous endonuclease, which cleaves chromosomal DNA at the junction of nucleosomes. Agarose gel electrophoresis shows a specific ladder pattern in apoptotic cells but a diffuse continuous pattern in dead cells.
Principle of the detection
This kit is based on TUNEL (TdT mediated dUTP Nick End Labeling) method. TdT (Terminal Deoxynucleotidyl Transferase) was used to catalyze the incorporation of FITC-12-dUTP at the 3′-OH terminus of the broken DNA in apoptotic cells. FITC-12-dUTP-labeled DNA can be directly observed by fluorescence microscopy or quantified by flow cytometry. This kit is optimized for the labeling reaction. It uses the best proportion of fluorescence marker and unlabeled dNTP for incorporation of the 3′-OH terminal nucleotides so that the ends of the same fragmented DNA fragment can form longer „marker tail”. The “marker tail” reduces the steric hindrance of labeling groups adjacently incorporated into dNTPs. It increases the number of fluorophores on each fragment and reduces the possibility of adjacent fluorophores’ aggregation and quenching. Thereby, the “marker tail” improves detection sensitivity and reduces nonspecific reactions.
