IMPROVED TECHNIQUE FOR INCREASED GRANULOCYTE RECOVERY FROM CANINE WHOLE-BLOOD SAMPLES BY COUNTERFLOW CENTRIFUGATION-ELUTRIATION .1. INVITRO ANALYSIS

Abstract

Granulocytes were isolated from 120 ml of canine whole blood by a modification in counterflow centrifugation-elutriation technique. The leukocytes were concentrated in a buffy coat fraction and diluted to a fixed RBC [erythrocyte] density prior to entry into the rotor. Overall recovery of granulocytes from canine whole blood averaged 82% with 96% purity from contaminating mononuclear leukocytes. The purified cellular fraction was assayed in vitro to monitor O2 consumption, chemotaxis, bacterial growth inhibition and enzyme activities. Cellular integrity was monitored by scanning and transmission electron microscopy as well as by cell volume analysis. The granulocytes isolated by counterflow centrifugation-elutriation apparently suffered no noticeable morphological damage or loss of function in terms of recognition of a toxic agent, migration, phagocytosis or bactericidal capacity. Granulocytes isolated by counterflow centrifugation-elutriation may be physiologically more active than cells obtained by filtration leukapheresis, continuous-flow centrifugal leukapheresis or discontinuous flow centrifugation in vitro and in vivo.