EVALUATION OF INTRACELLULAR KILLING OF BACTERIA BY ENRICHED POPULATIONS OF MOUSE PERITONEAL EXUDATE NEUTROPHILS

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Abstract
Elicited, mouse peritoneal exudate cells were fractionated by centrifugation on discontinuous Percoll density gradients. Two subpopulations of neutrophils, each of greater than 90% purity, were isolated at discontinuous density gradient interfaces different from the region of mononuclear cell enrichment (i.e., 1.0694-1.0871 and 1.0872-1.1002 g/ml for neutrophils and less than 1.0694 g/ml for mononuclear cells). Peritoneal exudate cells were mixed with Proteus mirabilis in the presence of 1% normal mouse serum for 30 min. The mixtures were fractionated on gradients of Percoll diluted with a clacium-free medium. Populations of cells banding at densities greater than 1.0693 g/ml were washed free of gradient material, and neutrophil suspensions containing intracellular bacteria and which were relatively free of extracellular bacteria were isolated. Less than 7% of the total bacteria present was extracellular. The continuing extracellular presence of a heat-labile component of normal mouse serum was essential for maximal intracellular kill of P. mirabilis by mouse peritoneal neutrophils.

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