Analysis of Cell Cycle Distribution of Human Bladder Cancer Cells by BrdU-Hoechst Flowcytometry

Abstract

The natural history and therapy of bladder tumors are dictated primarily by the stage and histopathology of the disease. Nevertheless, among this tumor type there is a broad spectrum of biological behavior, and its response to a particular treatment regimen is highly variable. One reason for this heterogeneous biological behavior could arise from differences in cell cycle kinetics. Traditionally, in vitro growth and cell cycle kinetics of bladder tumors have been investigated by mitotic chromosome labelling techniques, such as radioactive thymidine uptake (Morimoto et al. 1980) or BrdU-incorporation (Hefton et al. 1980). Except for an accurate but time-consuming double-labelling technique (Schultze 1981) these standard methods do not provide exact estimates of the growth fractions. Moreover, the identification of kinetically divergent subpopulations within the heterogenic bladder tumor cell culture has not been possible. The introduction of monoclonal BrdU antibodies have improved the estimates of S-phase cell fractions, but there is no evidence that this new technique can differentiate between phase fractions which belong to successive cell cycle generations. Here, we introduce an alternative technique which likewise uses the principle of BrdU-incorporation, but which detects BrdU-substitution by its quenching effect on Hoechst-dye fluorescence (Latt et al. 1977).