ISOLATION OF RADIOPURE PLASMA AND HEPATIC ALBUMIN IN ACUTE-PHASE CONDITIONS

It is essential to obtain biochemically and radioactively pure albumin in studies on albumin metabolism and kinetics in stress and nutrition related conditions. However, published work on albumin metabolism, in both animals and man with acute phase reactions has usually been base d on inadequate chemical methods for isolation of homogeneous albumin free from acute phase proteins and other contaminants. Applications of conventional antibody precipitating techniques was usually either not sufficient to give radiopure albumin, or did not allow determination of the true specific radioactivity during in vivo experiments. Thus, t he lack of applicable methods to achieve radiopure albumin from small plasma and tissue samples for subsequent analyses and determination of the true specific radioactivity in albumin initiated the present meth od development. The combination of HPLC ion-chromatography (DEAE-sepha rose), affinity chromatography (Blue sepharose CL-6B, Con A sepharose) and HPLC based size exclusion chromatography (Protein PAK 300 SW, Wat ers) was applied. By this procedure we obtained radiopure albumin from both plasma and hepatic samples from individual mice with acute phase response as confirmed by two-dimensional electrophoresis and immune p recipitation.