Department of Immunology and Infectious Biology, University of Lodz, Lodz, Poland
Background: The immunomagnetic separation technique is the basis of monocyte isolation and further generation of monocyte-derived dendritic cells. Objective: To compare the efficiency of monocyte positive and negative separation, concentration of beads, and their impact on generated dendritic cells. Methods: Monocytes were obtained using monoclonal antibody-coated magnetic beads followed the Ficoll-Paque gradient separation of mononuclear cell fraction from the peripheral blood of 6 healthy volunteers. CD14 expression was analyzed by flow cytometry. Results: The percentage of MDDCs generated from monocytes obtained by positive and negative selection was comparable (51.8 ± 15.0 and 46.7 ± 3.4, respectively; p=0.885). The median values for the number of MDDCs obtained from monocytes after positive selection (3.9 × 106) and for MDDCs obtained from monocytes after negative selection (3.1 × 106) were comparable (p=0.194). The use of the recommended or half of the amount of beads for both types of separation had no significant influence on the percentage of isolated cells. Conclusions: Both types of magnetic separation including recommended and reduced concentrations of beads did not affect the yield and the purity of monocytes and their surface CD14 expression. However, DCs originated from the “positively” separated monocytes had noticeable higher expression of CD80.