Crude bovine plasminogen, prepared from bovine serum by 30% saturation with ammonium sulphate, was partially purified by isoelectric precipitation at pH 5.3, then purified by chromatography on columns of diethylaminoethyl-cellulose. Additional purification of the column product was obtained by precipitation of plasminogen from solution with potassium phosphate. The most highly purified bovine plasminogen preparation, assaying 187 esterase units per mg nitrogen, represents a 28-fold purification of crude plasminogen. The over-all recovery from crude plasminogen was 15–20%. Sodium chloride inhibited crude plasminogen but had no effect on the activation of the most highly purified bovine plasminogen preparations. Bovine plasminogen could be spontaneously activated in 50% glycerol solution, but the degree of activation and the time required for maximum activation was also dependent on the purity of the preparation. Free boundary electrophoresis of purified preparations indicated that bovine plasminogen is associated with the major, faster-migrating component.