The FD Apop ™ kit is designed for the microscopic detection of cells undergoing apoptosis according to the principle of the in situ DNA notch end labeling (TUNEL) technique ¹.
The assay uses terminal deoxynucleotidyl transferase to catalyze the incorporation of biotinylated deoxyuridines at the free 3′-hydroxyl terminals of DNA fragments, which are considered one of the most characteristic features of apoptosis2, 3. Integrated biotins are amplified and visualized with the avidin- biotin complex (ABC) 4 method, which allows identification with light microscope.
The FD Apop ™ Kit reagents and procedure have been optimized to achieve a high degree of specificity and sensitivity for detecting apoptotic cells with minimal background. This kit can be used with frozen and paraffin sections, as well as with cultured cells. The kit procedure takes approximately 4 hours.
- Part I (Store at -20 ° C)
- Digestive enzyme 2 ml x 4
- Reaction solution A 2 ml x 2
- Reaction solution B 60 µl
- Reaction solution C 40 µl
- Chromogen solution 20 ml
- Part II (Store at 4 ° C)
- Balance buffer 20 ml
- Detection reagent 6 ml
- Phosphate Buffered Saline 10x 250 ml x 2
Materials required but not included:
- Bidistilled water
- Humidified chamber
- Incubator or water bath (30 ° C)
Histological supplies and equipment, including microscope slides, glass coverslips, staining bottles, fine tip forceps, ethanol, xylenes or xylene substitutes, mounting medium, and a light microscope.
Gavrieli Y, Sherman Y and Ben-Sasson SA. (1992) Identification of programmed cell death in situ by specific labeling of nuclear DNA fragmentation. J. Cell Biol. 119: 493-501.
Wyllie AH. (1980) Glucocorticoid-induced thymocyte apoptosis is associated with activation of endogenous endonucleases. Nature. 284: 555-6.
Arends MJ, Morris RG and Wyllie AH. (1990) Apoptosis: the role of endonuclease. Amer. J. Pathol. 136: 593-608.
Hsu SM, Raine L and Fanger H. (1981) Use of the avidin-biotin-peroxidase (ABC) complex in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures. J. Histochem. Cytochem. 29: 577-80.