Two-wavelength mercury arc lamp excitation for flow cytometric DNA-protein analyses

Correlated DNA- and protein-determinations were performed using flow cytometry and DAPI-sulforhodamine 101 (SR101) staining. Two-wavelength UV-green excitation using a single HBO lamp was found to be superior to single wavelength UV excitation, since it resulted in a higher specificity in the SR101 measurements. The effects of overlapping in the emission spectra and of energy transfer from DAPI to SR101 were minimised. Different staining conditions and the specificity of SR101 staining were examined. No binding of SR101 to DNA and RNA could be detected. Good reproducibility was achieved in the protein measurements using human lymphocytes as an internal reference. Examples are given of the analysis of mixed cell populations with different cellular protein contents and different growth and metabolic activities. Correlated measurements of DNA and protein appear to be useful for further analysis of tumor cell populations.