A pure lipase has been isolated from extracts of the human pancreas. The purification process includes centrifugation, two ion-exchange chromatography steps, and one gel filtration step. Compared with other reports, a high recovery, large amounts, and a high specific activity were obtained. Lipase is present at 1-2 mg/g in the pancreatic gland. In the absence of colipase and bile salts with tributyrine as substrate, the specific activity at room temperature and at pH 7.0 is 4000 mumol/min/mg. It increases to 8000-10,000 in the presence of colipase and bile salts at a temperature of 37 degrees C. The fate of the other human lipolytic proteins during the different purification steps is also indicated. Lipase purified by this method has been used for crystallization.