A rapid method for accurate DNA measurements in single cells in situ using a simple microfluorimeter and Hoechst 33258 as a quantitative fluorochrome.
A method is described for the rapid, accurate measurement of single cells in situ using microfluorimetry. This method involves a shutter system, which allows irradiation of single cells for fractions of a second and a peak fluorescence intensity recording device. In this way errors due to fluorochrome fading are almost eliminated and standard deviations of less than 5% are obtained. Hoechst 33258 has been used as a quantitative fluorochrome. Optimum fixation and staining conditions on glass and plastic tissue culture vessels are described.
D000818:Animals; D001562:Benzimidazoles; D006690:Bisbenzimidazole / Q000378:metabolism; D001854:Bone Marrow Cells; D003584:Cytological Techniques; D004247:DNA / Q000032:analysis* / Q000378:metabolism; D005404:Fixatives / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000166:cytology; D008214:Lymphocytes / Q000032:analysis; D051379:Mice; D012260:Ribonucleases / Q000494:pharmacology; D013050:Spectrometry, Fluorescence / Q000379:methods*
D001562:Benzimidazoles; D005404:Fixatives; D004247:DNA; D012260:Ribonucleases; D006690:Bisbenzimidazole