The effects of cell concentration and anticoagulants on neutrophil passive migration and chemotaxis were examined. A relationship between passive migration and cell numbers was documented using blood neutrophils from neutropenic and normal individuals, as well as with purified granulocytes suspended in autologous serum. At granulocyte levels below 1500/cu mm, passive migration is impaired. The apparent defect in neutropenic patients can be normalized by adjusting cell counts to normal levels. Inhibition of passive migration by standard concentrations of heparin and chelator anticoagulants necessitates the use of small and carefully controlled amounts of these agents. Cell concentration and anticoagulants likewise influence chemotaxis. EDTA, oxalate, and citrate prevent the formation of chemotactic factor(s), whereas heparin appears to inhibit chemotaxis by a different mechanism possibly related to a direct cell effect. A knowledge of these variables is essential for proper evaluation of leukocyte migration studies.