BIOCHEMICAL AND MORPHOLOGICAL CHARACTERIZATION OF AZUROPHIL AND SPECIFIC GRANULES OF HUMAN NEUTROPHILIC POLYMORPHONUCLEAR LEUKOCYTES

Abstract

Postnuclear supernates from homogenates of purified neutrophil polymorphonuclear leukocytes (PMNs) from human blood were fractionated by zonal sedimentation and isopycnic equilibration in sucrose gradients. The fractions were characterized biochemically by measuring protein content and the activities of eight enzymes. Selected fractions were further analyzed by electron microscopy. In both centrifugation systems, azurophil and specific granules could be resolved almost completely. Azurophil granules sediment three to four times faster than the specifics and have an average density of 1.23. They contain all the peroxidase of the cells, large portions of four lysosomal hydrolases, and about half of the total lysozyme, and therefore appear to be, in biochemical terms, very similar to the azurophil granules of rabbit PMNs. The specific granules, which have an average density of 1.19, contain the remaining half of the lysozyme but appear to be free of the other components of the azurophil granules, and of alkaline phosphatase. Isopycnic equilibration disclosed a minor lysosomal population, which strongly overlaps the specific granules, and made possible the identification of a membrane-fraction which is characterized by the presence of the thiol-sensitive acid 4-nitrophenyl phosphatase and of alkaline phosphatase.