Human eosinophil peroxidase: purification and characterization.
Human eosinophil peroxidase (EPO) was isolated from granules from granulocytes of a patient with hypereosinophilia. The granules were extracted by means of 0.2 M NaAc, pH 4.0. The purification steps included gel filtration chromatography on Sephadex G-75 superfine and ion-exchange chromatography on CM-Sephadex G-50. The purified protein showed one band on agarose-electrophoresis, a high peroxidase activity, and a 415-nm/280 nm ratio of 1.15. After reduction, EPO showed two bands on SDS-PAGE of m.w. 52,000 and 15,000, respectively. On gel filtration, the unreduced protein had a m.w. of approximately 77,000. Amino acid analyses showed a high content of arginine and aspartic acid. Monospecific antibodies to EPO were prepared in rabbits, and a specific radioimmunoassay was developed. There was an almost linear correlation between the content of EPO measured by the radioimmunoassay and the number of eosinophils in a mixed cell extract from reference material, indicating the eosinophil origin of EPO. The content of EPO was estimated to be 15.0 micrograms/10(6) eosinophils.
D000596:Amino Acids / Q000032:analysis; D000937:Antigen-Antibody Reactions; D002850:Chromatography, Gel; D047209:Eosinophil Peroxidase; D004802:Eosinophilia / Q000201:enzymology; D004804:Eosinophils / Q000201:enzymology*; D006801:Humans; D005779:Immunodiffusion; D007958:Leukocyte Count; D008970:Molecular Weight; D010544:Peroxidases / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D011863:Radioimmunoassay
D000596:Amino Acids; D047209:Eosinophil Peroxidase; D010544:Peroxidases