Picogram per cell determination of DNA by flow cytofluorometry

Abstract

Using nuclei isolated from less than 0.2 g tissue or 10(7) cells, a method is presented for the quantitative determination of amounts of DNA per cell at the picogram level. This technique is based on the enhanced fluorescence of 4',6-diamidino-2-phenylindole (DAPI) when it binds to DNA. A rapid, one-step nuclear isolation and DNA staining procedure is used to prepare tissue samples for flow cytometric analysis. Frozen tissues give results comparable to those for fresh tissue. Both chicken and trout erythrocyte nuclei were used as reference standards in the determination of amounts of DNA per diploid cell for several mammals and Amazon molly fish. The consistent values obtained for different tissues from the same organism show the accuracy of this method for DNA measurement.