Detection of oxidant producing-sites in glutaraldehyde-fixed human neutrophils and eosinophils stimulated with phorbol myristate acetate.

摘要: The aim of the present study was to detect oxidant-producing sites, and elucidate their dynamic reorganization in human polymorphonuclear leukocytes (PMNs) fixed with glutaraldehyde which preserves cell structure. In biochemical analyses, detectable O2− generation unfixed PMNs upon stimulation phorbol 12-myristate 13-acetate (PMA) presence cytochalasin B characterized by a lag period approximately 10 sec followed production. maximal rate reached 3.18±0.07 n mol/min/1×106 cells (mean±S.D.; n=4) after 30 stimulation. exposed PMA fixation generated without at 0.35±0.05 (mean±S.D.) addition NADPH as substrate suspension. cytochemical assays, we employed both B, then incubation reaction medium (pre-fixed cells) incubated containing (post-fixed cells). Oxidant pre-fixed detected FAD medium. No oxidant-reaction product seen stimulated for whereas oxidant visualized intracellular compartments 20 sec. fact that showed increased numbers structures compared those sec, demonstrates amount number increases between start PMA. These results using coincided findings obtained from assays B. observed elongated tubular were arranged radial fashion, associated plasma membrane PMNs, post-fixed exhibited slender spherical or rod-shaped various lengths. indicate can be elucidating sites PMNs.