A Comparison of Cell Separations Obtained with Centrifugal Elutriation and Sedimentation at Unit Gravity

Publisher Summary This chapter presents a comparison of cell separations with centrifugal elutriation and sedimentation at unit gravity. The elucidation of the mechanisms of cellular proliferation, differentiation, and function in living tissues is as much dependent on the availability of highly purified target cell preparations as it is on access to highly purified reagents, growth factors, and specific probes and on the capacity of the experimenter to rigidly control experimental conditions. These methods have been used to great advantage to identify, analyze, and resolve unique cell populations in complex tissues but are limited in preparative applications because of their inability to rapidly process large numbers of cells, particularly when large quantities of rare cells are required. Unit gravity sedimentation relies on the sedimentation of single cells through a nonlimiting density gradient in the earth's gravitational field. In elutriation, monodispersed cells sediment centrifugally, opposed by the centripetal flow of the suspending medium with the rate of sedimentation being determined by the sum of these forces. In theory elutriation enables larger numbers of cells to be processed more rapidly and efficiently than unit gravity sedimentation. Shearing forces and wall effects to which cells are subjected are potentially far more detrimental during elutriation than during centrifugation in tubes. Cells fanning out radially from the center of rotation in the centrifugal field strike the wall of the chamber and fail to move to their equilibrium position, impairing resolution. Under these conditions, cells will also tend to aggregate, leading to impaired recovery. Temperature fluctuations caused by periodicity in centrifugal temperature control systems have also been documented as a cause of problems unique to elutriation, affecting both recovery and resolution. The impaired resolution and reproducibility caused by temperature fluctuations in elutriation experiments described in the chapter are mediated by changes in the viscosity of the medium, which, in turn, affect the coefficient of variation of the sedimentation rate.