| In biomedical research and in the biotechnology industry, there have been increasing demands for a wide range of techniques that can perform high efficiency cell separation, as well as complex cellular analysis.; Among the major modern methods for cell separation and characterization, the steric field-flow fractionation (FFF) technique is a relatively new and powerful method capable of performing rapid and convenient separations at a low cost. The purpose of this study is to develop a better understanding of the separation mechanism of steric FFF, and to develop new applications of this technique in clinical diagnosis and cell separation.; In the first phase of this work, a systematic study using model particles is carried out to explore the separation mechanism of the steric FFF system. Various system parameters (carrier flow velocity, field strength, and carrier viscosity) and particle properties (size, shape, deformability) are found to be influential on particle retention behavior, and hence the resolving power of the technique.; The second phase of this work focuses on developing new applications of the steric FFF technique in clinical diagnosis of erythrocyte abnormalities. First, various chemical fixations of human erythrocytes are induced to mimic certain physiological conditions known to increase rigidity of red cells, and the steric FFF technique is shown to allow a differentiation of red cells with different deformability. A multilaboratory study then is conducted to provide a comparison of the performance of FFF systems, and channel configuration is shown to have minor effects on separation resolution. This technique is also used to examine red cells for potential effects of parenteral fat emulsions, and significant effects are noted, encouraging further investigation.; In the third phase of the study, FFF is being used to provide rapid separations of viable CHO-K1 cells and their high permeability mutants, based on differences in their swelling kinetics. This is the first attempt in FFF development to separate particles according to a nonequilibrium property. To generalize and to optimize this separation process, a numerical simulation is developed, and optimal operating parameters predicted by the computer model are in good agreement with available experimental data. |
