Purification of Fc gamma receptor from rabbit alveolar macrophages that retains ligand-binding activity.
The Fc gamma receptor of rabbit alveolar macrophages was purified by affinity chromatography by using rabbit gamma-globulin (Rab gamma G) coupled to Sepharose. Macrophage preparations were efficiently labeled with 125I by using a modified lactoperoxidase method. After incubation of NP-40 cell lysates with Rab gamma G-Sepharose, elution at 4 degrees C with 0.5 N acetic acid containing 1% NP-40 and rapid neutralization allowed recovery of active Fc gamma receptor. Purified Fc gamma receptor retained its ligand-binding activity, since approximately 41 to 72% of labeled material specifically rebound to Rab gamma G-Sepharose. Active receptor also rebound to human IgG- and rat IgG-sepharose. Active Fc gamma receptor did not bind to Sepharose coupled to rabbit Fab, rabbit F(ab)'2 or human F(ab)'2 fragments, nor to Sepharose coupled to chicken IgG. Analysis of Fc gamma receptor by SDS polyacrylamide gels demonstrated a broad peak of radioactivity in the apparent m.w. range of 50,000 to 70,000 in 5.6% acrylamide gels and 35,000 to 55,000 in 9% gels. Labeled receptor with similar structural characteristics and ligand-binding activity was also obtained from highly purified adherent cell populations and from macrophages biosynthetically-labeled with [14C]glucosamine in culture.
D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion; D002469:Cell Separation; D004591:Electrophoresis, Polyacrylamide Gel; D007164:Immunosorbents; D008024:Ligands / Q000494:pharmacology; D046911:Macromolecular Substances; D008264:Macrophages / Q000276:immunology*; D008970:Molecular Weight; D011817:Rabbits; D011961:Receptors, Fc / Q000096:biosynthesis / Q000302:isolation & purification*; D012685:Sepharose / Q000276:immunology; D012995:Solubility
D007164:Immunosorbents; D008024:Ligands; D046911:Macromolecular Substances; D011961:Receptors, Fc; D012685:Sepharose