A simple method for isolating human and rabbit polymorphonuclear neutrophils (PMNs)

We report the successful application to human venous blood of a novel metho d developed to purify rabbit polymorphonuclear neutrophils (PMNs) from whol e blood. Human PMNs were separated from whole blood after sedimentation wit h dextran and histopaque density gradient centrifugation, 3.92+/-0.26x10(6) PMNs per mi of blood was harvested. The purity of the preparation was 92.0 0 +/- 1.10%. The PMNs isolated were capable of generating a high amount of reactive oxygen species (ROS) and elastase after stimulation with phorbol 1 2-myristate 13-acetate (PMA): 13.43+/-0.3 muM of O-2(-), 9.62+/-0.15 muM of H2O2 and 5.48+/-0.01 muM of elastase. This method gives equivalent yield a nd viability when applied to isolating human or rabbit PMNs, in comparison with standard methods used to isolate human PMNs. Our method could be usefu lly exploited for comparative studies of rabbit and human PMNs with a cellu lar model of inflammatory oxidative stress in which the monitoring paramete rs are ROS and elastase, Thus, the results of animal (rabbit) studies tan b e extended to human diseases.