Murine monoclonal antibodies (mAb) from cell culture supernatants have been purified in order to acquire clinical grade for in vivo cancer treatment. The starting material was purified by high performance liquid chromatography (HPLC) systems ranging from the analytical scale process to a scaleup to 1 g per batch. Three columns (Protein A affinity chromatography with single-step elution, hydroxyapatite (HA) chromatography followed by linear gradient elution and endotoxin removing-gel chromatography), exploiting different properties of the mAb were applied. The final batches of antibody were subjected to a large panel of tests for the purpose of evaluating the efficacy of the downstream processing. The resulting data have allowed us to determine the maximum number of times the column can be used and to precisely and thoroughly characterize antibody integrity, specificity, and potency according to in-house reference standards. The optimized bioprocessing is rapid, efficient, and reproducible. Not less importantly, all the techniques applied are characterized by costs which are affordable to medium-sized laboratories. They represent the basis for implementing immunotherapeutic protocols transferable to clinical medicine.