Leukocyte Isolation by Sedimentation: The Effect of Rouleau-Promoting Agents on Leukocyte Differential Count

Abstract

The rouleau-promoting agents dextran and polyvinylpyrrolidone (PVP) were used to accelerate erythrocyte sedimentation in order to harvest the leukocyte rich plasma (LRP). The objective of the work was to determine if agent concentration or blood: agent ratio had any effect on the leukocyte differential count and if so at what agent concentration and agent:blood ratio did the LRP leukocyte differential count most closely match the whole blood leukocyte differential count. With both sedimentation agents the data clearly indicate that both parameters effect LRP differential counts and that low concentrations of sedimentation agents are most important in obtaining LRP differential counts which most closely match the whole blood differential counts.

A number of methods have been used to isolate leukocytes from whole blood, including the selective destruction of erythrocytes, sedimentation, simple centrifugation, leukoad-hesion, use of phagocytic properties and density gradient methods^{2, 3}.

One of the simplest techniques is to use a rouleau-promoting agent to accelerate the sedimentation of the erythrocytes, followed by the harvesting of the supernatant plasma or leukocyte rich plasma (LRP). The LRP may be used as collected or, with centrifugation, a concentrate of leukocytes may be obtained. A variety of sedimentation-enhancing materials have been used, including fibrinogen^{4, 10}, dextran, phytohemagglu-tinin¹¹, polyvinylpyrrolidone (PVP), gelatin¹², gum acacia¹³and hydroxyethyl starchl^{14, 15}. The most frequently used of these appear to be dextran and PVP^{2, 3}. Many papers describe the use of these agents with several reporting results of various procedures on total leukocyte yield and erythrocyte contamination^4–8 or enhanced granulocyte yield^{14, 15}. The relationship between LRP differential count and the whole blood differential count with the use of these agents at various concentrations and blood:agent volume ratios has yet to be reported and is the subject of this communication.