Isolation and Phagocytic Properties of Neutrophils and Other Phagocytes from Nonmastitic Bovine Milk

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Abstract
A technique for the separation of neutrophils from macrophages-epithelial cells in samples of nonmastitic bovine milk with low cell counts has been developed. The procedure is based on centrifugation in a discontinuous metrizamide gradient and is rapid, taking less than 40 min. The recovery of the neutrophils is about 30% and their viability about 90%. The isolated neutrophils showed an appreciable unstimulated luminol- and lucigenin-dependent chemiluminescence, which was due to NADPH oxidase rather than to xanthine oxidase. The neutrophils had a higher rate of ingestion of C3-opsonized particles than macrophages-epithelial cells, whereas no significant differences in phagocytosis of IgG-opsonized yeast or unopsonized yeast were detected between the two cell populations. The macrophages-epithelial cells produced no luminol-dependent chemiluminescence and induced considerably lower activity in the lucigenin-dependent system than neutrophils, indicating that these cells contain no myeloperoxidase. Analyses of the activity of the neutrophils in response to C3-opsonized yeast particles showed that the luminol-dependent chemiluminescence of cells isolated from residual milk increased significantly over the lactation period. Moreover, a tendency to a higher phagocytosis and chemiluminescence of neutrophils isolated from residual milk than from stripping milk was indicated.

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