Analysis of minor proteins in animal sera is of considerable clinical significance. To be able to detect these proteins, depletion of major proteins (albumin and immunoglobulin G [IgG]) is necessary. Many of these proteins are also required in pure form for a variety of biochemical applications. The present work uses goat serum as the system and describes the separation and purification of both major and several minor proteins. This was carried out by judicious adaptation and combination of separation technologies such as immobilized metal ion affinity chromatography (on a somewhat novel matrix), dye affinity chromatography, and lectin affinity chromatography. Albumin, IgG, α(2)-macroglobulin, α(1)-proteinase inhibitor, and transferrin were obtained from the serum. The purified preparations were found to be homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis.