Dynamic cellular events

Abstract

Measurement of dynamic cellular events in intact cells under near physiological conditions, irrespective of the assay system, is arguably one of the most powerful techniques in cell biology, as physiological and pathological processes are not static but are continuously variable. It is becoming increasingly obvious that many of the initial events involved in cell proliferation, “activation” of all types and differentiation control take place within a time frame of seconds, but the ultimate effects of those initiating events (e.g. growth-factor stimulation, membrane calcium flux, pH changes and carcinogen interaction) may only become apparent some days, weeks or even years later. Clearly, an understanding of the dynamics of inappropriate or abnormal initiating events will be of crucial importance in attempts to modulate pathological states. This chapter addresses flow cytometric analysis procedures to measure dynamic events. Incorporation of time Incorporation of time in the data base is the most important single feature in the measurement of dynamic cellular processes using flow cytometry, and a number of different techniques have been implemented for this. Watson et al. (1977) used a continuous interrupted sampling technique, where the cells were continuously flowing through the instrument. The data-acquisition computer was instructed to record for 5 seconds and then wait for 10 seconds before re-commencing acquisition. During the interval between recordings the median of the population was calculated and printed out.