Fluorometric determination of DNA in epidermis and cultured fibroblasts, using 4′-6-diamidino-2-phenylindole (DAPI)

✍ Scribed by J. Ch. Meyer; H. Grundmann

• Book ID: 104763757
• Publisher: Springer-Verlag
• Year: 1984
• Tongue: English
• Weight: 372 KB
• Volume: 276
• Category: Article
• ISSN: 0340-3696
• DOI: 10.1007/bf00412563

✦ Synopsis

4'-6-diamidino-phenylindole (DAPI) which is used for the detection of mycoplasmic infections of cell cultures or cytofluorometric investigations proved to be a sensitive agent for the fluorometric assay of alkali extracted DNA in cultured cells and human epidermis. Various experiments to optimize assay conditions led to the following procedure: 1 part of freeze dried tissue (1 mg/ml) and I part of 0.2 tool/1 NaOH (w/v) were heated at 95 ~ for 40 rain. After cooling for 1 h to room temperature, 0.1 ml alkaline extract was mixed with 2.9ml DAPI-solution (0.2gg/ml in McIlvaine buffer containing I mmol/1 MgC12). After 10 min in the dark the emitted fluorescence was estimated at 453 nm with an excitation at 360nm. This assay procedure allowed a detection limit of 25 ng DNA.