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Quantitative Analysis of the Cell Cycle in Imaging Applications

Introduction

Quantitative analysis of stages in the cell cycle is commonly used in cell biology since many pharmacological interventions aim to modulate the cell cycle in some way. We present here an application that can be used in adherent cells and optionally multiplexed for the use in automated cell signaling analysis and drug-screening applications.

The cell cycle consists of a series of processes involved in the growth, replication, and division of cells and can be divided into two major stages, interphase (the phase between mitotic events) and mitosis. There are three distinct, successive stages within interphase, namely G1, S, and G2 phases. During G1 (first gap), cells monitor their environment and grow (synthesize RNA and proteins). Once conditions are optimal, cells commit to DNA synthesis (S phase) and replicate their chromosomal DNA. The G2 phase (second gap) follows when cells continue to grow and prepare for mitosis (M phase, division). The G2 gap allows time for the cell to complete its DNA replication before initiating mitosis.

During mitosis, the cell cycle is completed giving rise to two daughter cells, each of which contains the same genetic material as the original cell and approximately half of its G2 level of cytoplasm. In addition to these specific stages, a G0 phase has been described for cells that exit from the cellcycle and enter a quiescent, non-dividing state. In response to external stimuli, cells may undergo reactivation and leave G0 and enter the G1 phase of the cell-cycle. Another consequence of cellular activation may be the induction of programmed cell death (apoptosis).

Understanding the specific phase of a cell is important when cells display heterogeneous responses to drugs as is often the case in high-content applications.

This four color application demonstrates how fluorescent antibodies can be used in comb
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