Comparison of three monocyte depletion techniques for lymphocyte isolation from peripheral blood mononuclear cell

Abstract

Background: Lymphocytes are crucial cells in the immune system. Studying lymphocyte function can lead to better understanding of the immune system. Essentially, lymphocyte isolation technique is required for studying lymphocyte function. Several techniques were developed to prepare lymphocytes, including depletion of monocytes from peripheral blood mononuclear cells (PBMCs). Objectives: To compare utilization of three different techniques for lymphocyte isolation by monocyte depletion from PBMCs. Materials and methods: Lymphocytes were isolated from PBMCs by depletion of monocytes using (i) magnetic beads phagocytosis, (ii) Percoll density gradient centrifugation, and (iii) anti-FITC antibody conjugated micro-magnetic beads. The number of cells collected was counted using Turk’s solution. The cellular profiles of PBMCs and monocyte-depleted PBMCs were determined by immunofluorescence and flow cytometry. Results: The highest yield and purity of monocyte-depleted PBMCs were achieved using the anti-FITC antibody conjugated microbeads depletion method. However, this method consumed the longest time and had the highest cost. Magnetic beads phagocytosis depletion method required the shortest time; however, the wide range of collected yield was a concern. Percoll gradient centrifugation method was the cheapest, but the percentage yield was the lowest among the three methods. Conclusion: The utilization of three different methods was able to deplete monocytes from PBMCs. However, each technique had some advantages and disadvantages. The information obtained from this study might give some guidance for selecting a suitable method for isolation of lymphocytes based on the monocyte-depleted PBMC strategy.