Analysis of Velocity Sedimentation Techniques in Cell Separation. Influence of Apparative and Sample Properties on Separative Power, Resolution and Sensitivity

Abstract

Summary: In the present investigation the velocity sedimentation technique was analysed with respect to separation resolution, power and sensitivity. It was found that apparative modifications do not influence the resolution, which is a function of the contribution of apparative errors to the dispersion. A surprisingly small parameter °f 0.15 was determined and it seems unlikely that this value can be improved. On the other hand an a Pparative modification is presented which improves the separation power and makes sample loading independent of the gradient filling. If cells (from rat bone marrow) were separated, a several times higher dispersion for a given cell volume was observed than was due to the apparative error. It was concluded that density variations were the major source of this dispersion. Since cell volume and density apparently show independent variations within a biological cell population the cell density cannot be disregarded if velocity sedimentation profiles are discussed in physical terms as is often done.

Synchronization of mammalian cells by selection and additional chemical block studied by DNA distribution analysis and BrdUrd‐Hoechst 33258‐technique

“…An analysis of the selected subpopulations, including measurements of cell number, DNA distributions and volume distributions has not only shown the general applicability of the zonal rotor method for selecting synchronous cells, but also the limitations of this method. Separation of partially synchronous cells in a zonal rotor has been studied earlier (Probst & Meisenbacher, 1973;Warmsley 8c Pasternak, 1970) with similar results, but the limitations of this method have not been investigated in detail, as has been done for separation of cells by a l g separation chamber (Zeiller et al, 1976). The main reason for the limited separation of cells using zonal centrifugation was the design of the zonal rotor.…”