Cell-Cycle Analysis in Phagotrophic Microorganisms from Flow Cytometric Histograms

The inability of certain phagotrophic protozoa to phagocytose right before and right after cell division was utilized to measure the distributions of newborn and dividing cells and obtain information on the kinetics of the cell cycle. A fluorescent microsphere labeling assay was used in conjunction with flow cytometry to identify the non-feeding cells and measure their DNA and protein distributions. These distributions, which are typically bimodal, were decomposed to the two component distributions by means of a new algorithm, which is based on the assumption that cells partition their DNA and the remaining cellular material evenly at division. The estimated population fractions in the two non-feeding phases before and after division and the corresponding DNA and protein distributions provide significant information on the kinetics of the cell cycle. The transient labeling experiments revealed a significant variability in the timing of the pre-division non-feeding phase with the smaller cells of this subpopulation spending more time in preparation for division. From the determined distributions of newborn and dividing cells, the single-cell growth rates and division probabilities were also calculated. This type of analysis provides essential information for the development of corpuscular, multi-staged cell population models.