Separation of human granulocytes and mononuclear cells from whole blood using percoll on a centrifugal microfluidic disc

Abstract

In this study, a new and robust strategy based on a lab-on-a-disc (LOD) platform for isolating human white blood cells (WBCs) from whole blood is presented. Fully automated disc performance is defined based on standard cell separation techniques with density gradient media (DGM). In this centrifugal microfluidic system, capillary and siphon valves' integration controls liquids' flow and handling. The one-step isolation process performed depends on the selective sedimentation on the percoll and no need for labeling. This pattern is achieved without any additional steps of extracellular filtration or cell lysis by optimizing the concentration of percoll media and centrifuge conditions. Five percoll media in the density range of 1.058–1.095 g.ml−1 have made it possible to achieve a cell band with the desired recovery rate and purity. Sedimentation process performed using gradient media, 1.058–1.077, and 1.077–1.088 g.ml−1 at a purity rate of 83% and 96%, respectively, in mononuclear cells (MNCs) and neutrophils isolation has recorded. For having high purity in blood cell analysis, a gradient medium with a density of 1.088 instead of 1.095 g.ml−1 has been used. Thus, the level of red blood cells (RBCs) contamination in the isolated granulocyte layer is effectively reduced. The obtained data prove that cellular viability is maintained during the performed processes, and the number of dead cells is less than 10%. Therefore, the proposed microfluidic disc is the right candidate in a wide range of bioanalysis assays and clinical diagnostics with a meager amount of sample.