An enzyme with proteolytic activity has been isolated from the subcellular granules of rabbit polymorphonuclear leucocytes. Purification of the enzyme involved extraction of the granule membranes with 0.01 M sodium phosphate buffer, pH 7.4, containing 1 M NaCl and 0.1% Triton X-100, followed by gel exclusion chromatography on Sephadex G-75. The enzyme hydrolysed p-nitrophenol-N-tert-butyloxylcarbonyl-L-alaninate, a synthetic substrate for elastase, but failed to hydrolyse elastin. The enzyme also hydrolysed azo-albumin with a pH optimum between 7.5 and 8.5. Inhibition studies indicated that the enzyme was a serine proteinase (EC 3.4.21.-) and it was found to have an apparent molecular weight of 25 000 by polyacrylamide gel electrophoresis. The purified enzyme behaved as a single on SDS-polyacrylamide gel electrophoresis, had a single isolectric point at pH 5.9, yet showed multiple components on centrifugation.