Platelets augment granulocyte aggregation and cytotoxicity: uncovering of their effects by improved cell separation techniques using Percoll gradients

The ‘standard’ technique of granulocyte preparation for in vitro studies uses dextran removal of erythrocytes and Ficoll‐Hypaque gradient centrifugation to increase granulocyte purity. The procedure is lengthy, approximately 150 min in our hands, and provides granulocytes significantly contaminated with platelets (approx. 5 platelets/PMN). We report a technique that replaces dextran with hydroxy‐ethylstarch and Ficoll‐Hypaque with Percoll. Preparation time is reduced by approximately 40% and platelet contamination by more than 80%. Granulocytes, so prepared, function metabolically (O₂‐generation, chemiluminescence, HMP‐shunt maxima) and, in motility/phagocytosis assays, identically to ‘standard’ preparations. However, an augmentatory effect of platelets in granulocyte aggregation responses and their mediation of cytotoxicity is uncovered. Ficoll‐Hypaque purified cells (platelet‐rich) aggregate to a significantly greater degree with FMLP or activated complement lectins and excessively kill ⁵¹Cr‐labelled target cells when compared to Percoll‐preparations (platelet‐poor). Re‐addition of purified platelets or of platelet release supernatants to the latter reproduces results using the ‘standard’ preparations.