Chapter 3 - Sedimentation of Cells: An Overview and Discussion of Artifacts

Abstract

This chapter discusses sedimentation at unit gravity, elutriation, isokinetic sedimentation, and the separation of cells by velocity sedimentation in a reorienting gradient zonal rotor. The term velocity sedimentation refers to the kind of sedimentation that occurs before cells arrive at their respective buoyant densities. For most kinds of velocity sedimentation, it is desirable to use gradients with densities as remote as possible from the densities of the cells. For maximal resolution, these gradients should increase in the concentration of solute. There are four forms of velocity sedimentation that are broadly applicable to the separation of viable cells: sedimentation at unit gravity, sedimentation in an isokinetic gradient of Ficoll in tissue culture medium, elutriation or counter-streaming centrifugation, and centrifugation in a reorienting gradient zonal rotor. Sedimentation at unit gravity and isokinetic sedimentation offer the investigator two major advantages: (1) Cells can be separated under sterile conditions with a high degree of reliability and (2) the necessary equipment is either available in most laboratories or very inexpensive. Both techniques are best suited for work with relatively small numbers of cells. In contrast, elutriation and sedimentation in a reorienting zonal rotor require much more skill and care if they are to be used for work under sterile conditions. Isopycnic or buoyant density sedimentation refers to the sedimentation of cells in continuous gradients with sufficient force and for a sufficient period of time for them to arrive at the locations of their respective densities in the gradient.