Density gradient ultracentrifugation of lipoproteins using metal ion complex solutes.

摘要

A lipoprotein profile measures the distribution of lipoproteins in serum. Since many of the risk factors for cardiovascular heart disease are associated with the serum lipoproteins, these profiles are important tools for assessing risk. The most common method for generating a profile is by separating the lipoproteins in a density gradient. Many descriptions of lipoprotein density profiling techniques can be found in literature. Most of these methods require very long centrifugation times. The more rapid methods suffer from poor resolution or they do not provide valuable density information.; The research presented in this dissertation describes an improved method for generating lipoprotein density profiles. This method consists of prestaining serum with a fluorescent probe followed by an isopycnic separation of the lipoproteins using a novel gradient. The solute responsible for generating this gradient is a metal ion complex. The fluorescence of the separated lipoproteins was measured by digital imaging.; This novel gradient, combined with the use of a fluorescent stain and digital imaging has maximized the amount of information that can be obtained from a lipoprotein density profile. Using this method, it was possible to determine the equilibrium densities of the lipoproteins as well as their concentrations from an image of a centrifuge tube. The entire process, including staining and analysis, requires less than six hours.