Flow Cytometric DNA Analysis in Oncology: From Single to Multiparametric Measurements/Durchflusszytometrische DNA-Analytik in der Onkologie: Von einzel- zu multiparametrischen Messungen

Abstract

DNA analysis was one of the earliest applications in flow cytometry. The stoichiometric interaction of fluorescent dyes with nucleic acid allows the evaluation of DNA content (ploidy) and the proliferative fraction. Initially, DNA was measured as the only parameter along with side-and forward-scatter properties to identify intact nuclei or cells. Later, a dual-parametric approach for the simultaneous analysis of DNA and protein was introduced. Today, the main application of flow cytometric DNA analysis is the additional, tumour entity-dependent diagnosis in pathology. The knowledge about an abnormal DNA content as a crude indicator of chromosomal instability and high S-phase fraction can result in a precise determination of tumour prognosis and can lead to the optimization of individual therapy and complementary disease monitoring. DNA analysis is applied in both haematological and solid tumour diagnosis. In contrast to haematological diseases, the analysis of solid tumours requires the isolation of cell nuclei or cell disaggregation with preservation of cell integrity. The uniparametric investigation of cell nuclei in suspension allows high-resolution DNA analysis with detection of aneuploidy in the neardiploid range. However, the multiparametric approach on the basis of cell selection or discrimination takes tumour heterogeneity into account, and DNA assessment can be combined with antigen detection with additional cell phenotyping. In contrast to unimodal DNA analysis, the multiparametric approach allows a more accurate calculation of cell-cycle fractions of individual cell subpopulations. All approaches require adequate standardization regarding cell preparation, data acquisition and handling to ensure lab-independent DNA data interpretation and utilization.