The fragmentation of genomic DNA by cellular nucleases during the later stages of apoptosis is also one of the most easily measured features of apoptotic cells. Nuclease activity generates DNA fragments ranging from ~300 bp to 50 bp in length, resulting in a typical DNA ‘laddering’ appearance when analyzed by agarose gel electrophoresis. These fragments have exposed 3’-hydroxyl (OH) ends which can be labeled with deoxyuridine triphosphates (dUTP). An enzyme, terminal deoxynucleotidyl transferase (TdT), is used to catalyze the template-independent addition of fluorescent tagged dUTP to the 3’-OH ends of double or single stranded DNA. This method is often called TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) or end labeling.
With the APO-DIRECT™ Kit, cells are stained with FITC-labeled dUTP in a single step. Samples can then be analyzed via flow cytometry. Samples that are apoptotic will stain brightly due to the substantial number of exposed 3’-OH sites, while cells that are non-apoptotic will not have incorporated significant amounts of FITC-dUTP and will stain dimly.
The APO-DIRECT Kit is shipped in one container and consists of two packages plus instruction manual. Upon arrival one should be stored at 2-8°C and the other at -20°C.
Lee KS, Lee J, Lee P, Jeon BC, Song MY, Kwak S, Lee J, Kim JS, Kim DJ, Kim JH, Tesh VL, Lee MS, Park SK. Inhibition of O-GlcNAcylation protects from Shiga toxin-mediated cell injury and lethality in host. EMBO Mol Med. 2022 Jan 11;14(1):e14678. doi: 10.15252/emmm.202114678. Epub 2021 Nov 29. PMID: 34842355; PMCID: PMC8749473.