Enrichment of Cells in Different Phases of the Cell Cycle by Centrifugal Elutriation

Abstract

Understanding the molecular and biochemical basis of cellular functions involved in growth and proliferation requires the investigation of regulatory events that most often occur in a cell cycle phase-dependent fashion. Studies involving cell cycle regulatory mechanisms and progression invariably require cell cycle synchronization of cell populations. Several methods are employed for obtaining and examining synchronized cells as they pass through one or more rounds of the cell cycle. Most of these methods involve pharmacological agents that act at various points throughout the cell cycle. Because of adverse cellular perturbations resulting from many of the synchronizing drugs used, other synchrony methods, such as serum deprivation and contact inhibition, have been exploited. Although such procedures allow synchronization of cells in a particular phase of the cell cycle, these approaches do not allow enrichment of cells, simultaneously in various phases of the cell cycle, from exponentially growing cell populations. Centrifugal elutriation described for the first time by Lindahl (1) is used to enrich cells in different phases of the cell cycle simultaneously with minimum changes in conditions during cell culture. Centrifugal elutriation can be used to obtain samples of uniformly sized cells, and because cell size is correlated with cell cycle stage, these cells are synchronized with respect to their position in the cycle.