ApopTag® Peroxidase in situ detection kits label apoptotic cells in research samples by modifying DNA fragments utilizing terminal deoxynucleotidyl transferase (TdT) for detection of apoptotic cells by immuno-peroxidase detection of digoxigenin-labeled genomic DNA in thin sections of fixed tissue.
ApopTag® Peroxidase in situ detection kits stains DNA strand breaks which in turn is detected by the TUNEL assay and subsequently visualised by light microscopy.
TUNEL kit for use with light microscopy ApopTag™ Peroxidase in situ Detection Kits detect apoptotic cells by immuno-peroxidase detection of digoxigenin-labeled genomic DNA in thin sections of fixed tissue. The ApopTag™ Peroxidase Plus kit includes positive control slides to aid identification of apoptotic cells. Principles of the procedure: Apoptotic cells are detected in situ by specific end labeling and detection of the DNA fragments produced by the apoptotic process. Residues of digoxigenin-nucleotide are catalytically added to the DNA by terminal deoxyribonucleotidyl transferase (TdT), an enzyme which catalyzes a template-independent addition of deoxyribonucleotide triphosphate to the 3’-OH ends of double- or single-stranded DNA. The incorporated nucleotides form a random heteropolymer of digoxigenin-11-dUTP and dATP, in a ratio that has been optimized for antidigoxigenin antibody binding. The anti-digoxigenin antibody fragment carries a conjugated reporter enzyme (peroxidase) to the reaction site. The localized peroxidase enzyme then catalytically generates an intense signal from a chromogenic substrate. This system allows for sensitive and specific staining of the very high concentration of 3’-OH ends that are localized in apoptotic bodies.
Principles of the procedure: Apoptotic cells are detected in situ by specific end labelling and detection of the DNA fragments produced by the apoptotic process. Residues of digoxigenin-nucleotide are catalytically added to the DNA by terminal deoxyribonucleotidyl transferase (TdT), an enzyme which catalyzes a template-independent addition of deoxyribonucleotide triphosphate to the 3'-OH ends of double or single-stranded DNA. The incoorporated nucleotides form a random heteropolymer of digoxigenin-11-dUTP and dATP, in a ratio that has been optimized for antidigoxigenin antibody binding. The anti-digoxigenin antibody fragment carries a conjugated reporter enzyme (peroxidase) to the reaction site. The localized peroxidase enzyme then catalytically generates an intense signal from a chromogenic substrate. This system allows for sensitive and specific staining of the very high concentration of 3'-OH ends that are localized in apoptotic bodies.
Key Applications: Apoptosis Detection
Application Areas: Cell Viability Analysis; Cellular Imaging; Cell Structure Analysis
Research Areas: DNA Fragmentation; OTHER - Drug Discovery; Cell Viability & Cytotoxicity; Apoptosis
Product Type: Kits & Assays
Kit or Assay Type: Apoptosis Assay Kits
Components: Equilibration Buffer (S7105), Reaction Buffer (S7106), TdT Enzyme (S7107), Stop/Wash Buffer (S7108), Plastic Coverslips and Anti-digoxigenin-peroxidase, ApopTag™ positive control slides, DAB Substrate, DAB Dilution Buffer, affinity purified sheep polyclonal antibody
Detection Method: Chromogenic
Storage & Handling: Store the kit at -15°C to -25°C until the first use. After the first use, if the kit will be used within three months, store the TdT Enzyme at -15°C to -25°C and store the remaining components at 2°C to 8°C.