title: Down-regulation of L-selectin surface expression by various leukocyte isolation procedures.
abstract: L-selectin, a cell surface glycoprotein expressed on lymphocytes, granulocytes, and monocytes, has been implicated in lymphocyte homing and extravasation of phagocytic leukocytes into areas of inflammation. Considerable differences of L-selectin expression among various individuals has been reported, with clinical correlations to perinatal events, maturation, and circadian rhythm. In this study, L-selectin expression of various white blood cells was found to be differentially sensitive to ficoll-hypaque or percoll density gradient centrifugation. After density gradient centrifugation, a significant loss of median monocyte L-selectin expression was observed when compared to time and temperature-matched controls or results obtained by whole blood incubation with anti-L-selectin monoclonal antibodies followed by simultaneous leukocyte fixation and red cell lysis. Mock treatment itself was associated with a variable L-selectin loss of monocytes but not lymphocytes or granulocytes. Ficoll-hypaque or percoll density gradient centrifugation resulted in significant L-selectin down-regulation of lymphocytes while granulocytes separated from lymphocytes and monocytes by ficoll-hypaque or percoll retained full L-selectin surface reactivity. L-selectin downregulation was seen also after colloid sedimentation with hydroxy-ethyl starch. It is concluded that unseparated blood should be used for measuring L-selectin expression.
authors.affiliation: Department of Neonatology, KAVH Children's Hospital, Rudolf Virchow Medical Center, Free University, Berlin, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Stibenz
authors.forename: C
authors.initials: C
authors.lastname: Bührer
mesh_terms: D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002469:Cell Separation / Q000379:methods*; D002499:Centrifugation, Density Gradient; D015536:Down-Regulation; D005362:Ficoll; D006098:Granulocytes / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D019041:L-Selectin; D007962:Leukocytes / Q000378:metabolism*; D008214:Lymphocytes / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D009000:Monocytes / Q000378:metabolism; D011205:Povidone; D016201:Receptors, Lymphocyte Homing / Q000378:metabolism; D012822:Silicon Dioxide
chemical_list: D015815:Cell Adhesion Molecules; D008562:Membrane Glycoproteins; D016201:Receptors, Lymphocyte Homing; D019041:L-Selectin; D005362:Ficoll; C016039:Percoll; D012822:Silicon Dioxide; D011205:Povidone