Objective: To determine the optimal method for separating neutrophils for studying neutrophil polarization.
Methods: Human neutrophil was separated from healthy human peripheral blood by Percoll density gradient centrifugation and Dextran sedimentation. The cell polarization, purity and activity of the neutrophils were determined, and F-actin polymerization and [Ca2+]i were analyzed.
Results: No significant difference was found in cell polarization, purity and activity of the human neutrophils separated by Dextran sedimentation and Percoll density gradient centrifugation (P>0.05), but F-actin polymerization was inhibited in PMNs separated by Dextran sedimentation, and the peak value of [Ca2+]i was decreased by 25% in PMNs separated by Dextran sedimentation compared to the cells separated by Percoll density gradient centrifugation.
Conclusions: Both Percoll density gradient centrifugation and Dextran sedimentation can be used for isolating human neutrophils to study cell polarization, but the former method allows better isolation. Dextran sedimentation can be considered when a large number of neutrophils need to be separated.
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