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

Arch Dermatol Res. 1984;276(1):52-6. doi: 10.1007/BF00412563.

Abstract

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 1 part of 0.2 mol/l NaOH(w/v) were heated at 95 degrees for 40 min. After cooling for 1 h to room temperature, 0.1 ml alkaline extract was mixed with 2.9 ml DAPI-solution (0.2 microgram/ml in McIlvaine buffer containing 1 mmol/l MgCl2). After 10 min in the dark the emitted fluorescence was estimated at 453 nm with an excitation at 360 nm. This assay procedure allowed a detection limit of 25 ng DNA.

Publication types

  • Comparative Study

MeSH terms

  • Cells, Cultured
  • Cold Temperature
  • DNA / analysis*
  • DNA / isolation & purification
  • Epidermis / analysis*
  • Ethidium
  • Fibroblasts / analysis*
  • Fluorescent Dyes*
  • Humans
  • Indoles*
  • Spectrometry, Fluorescence

Substances

  • Fluorescent Dyes
  • Indoles
  • DAPI
  • DNA
  • Ethidium