Stimulatory effect of counterflow centrifugal elutriation in large-scale separation of peripheral blood monocytes can be reversed by storing the cells at 37 degrees C

Abstract

Transfusion of peripheral blood monocytes may be of benefit as adjuvant treatment of leukopenic patients with serious infections. To study the feasibility of this approach, a large-scale monocyte separation procedure employing leukapheresis, density gradient centrifugation, and counterflow centrifugal elutriation was established. By processing 5 to 6 liters of normal donor blood, it was possible to obtain a mean of 1.1 x 10(9) (range: 0.5-1.7 x 10(9) cells) of mononuclear cells, of which 89% (range: 82-94%) were monocytes by Wright's stain morphology. When the elutriation was performed in XVIVO-10, a commercially available, serum-free medium developed for adoptive immunotherapy, spontaneous secretion of superoxide by the monocytes was significantly higher than for monocytes elutriated in Hanks' balanced salt solution without calcium and magnesium or non-elutriated peripheral blood mononuclear cells. This stimulated state of the monocytes was observed both immediately after elutriation and after overnight storage at 4 degrees C, and it was not affected by the type of storage vessel used. Overnight storage of the monocytes at 37 degrees C resulted in a reversal of the stimulated state of the cells. Monocytes elutriated in XVIVO-10 and kept overnight at 4 degrees C released high amounts of arachidonic acid. A subsequent decrease in this release was seen after additional storage at 37 degrees C for 18 hours. These observations demonstrate that separation and storage variables have important effects on the state of stimulation of monocytes. Further investigations of such variables may suggest improved procedures for preparation and storage of these cells, as well as possible ways to stimulate monocytes prior to transfusion.