Pt-staining of peroxidatic reaction products at the ultrastructural level.
The use of H2PtCl6 is proposed for the selective visualization of the poly-DAB reaction product created, in aldehyde-fixed tissue, with the cytochemical reaction according to Graham and Karnovsky (1966) or to Hoefsmit (1975). At sites known to contain peroxidatic activity, at the ultrastructural level, an electron-dense reaction product is acquired in otherwise unstained ultrathin sections. The presence of the element platinum in these sites has been demonstrated by X-ray microanalysis, for both the endogenous peroxidase and peroxidase conjugated to antibodies. The absolute platinum concentration has been established in erythrocytes and the granules in eosinophils and monocytes by co-embedded, Pt-containing Chelex ion-exchange beads next to the cells. By the application of the method of integrated morphometrical and chemical analysis (de Bruijn and Zeelen 1984; de Bruijn 1985; de Bruijn and Cleton 1985), both the elemental concentration and the area occupied have been calculated for eosinophil granules. The mean Pt net-intensity values of the cytoplasmic areas, known not to contain the enzyme peroxidase has been measured, and compared to the mean net-intensity Pt values of the granules. It was noted that the cytoplasmic Pt net-intensity values were not zero. The two sets of values are expressed as a mean Pt granule/cytoplasm ratio, this ratio creates a value for the "selectivity" of the reaction. The application of a postfixation reaction with OsO4- containing media, at pH 7.4, in addition to the H2PtCl6 reaction, resulted in a contrasted poly-DAB reaction product at all sites known to contain peroxidatic activity. However, X-ray microanalysis revealed that in addition to platinum, osmium was present.(ABSTRACT TRUNCATED AT 250 WORDS)
D015100:3,3'-Diaminobenzidine; D000818:Animals; D004577:Electron Probe Microanalysis; D004804:Eosinophils / Q000201:enzymology* / Q000648:ultrastructure; D006651:Histocytochemistry / Q000379:methods; D008297:Male; D051379:Mice; D008854:Microscopy, Electron / Q000379:methods; D008954:Models, Biological; D009000:Monocytes / Q000201:enzymology* / Q000648:ultrastructure; D010529:Peritoneal Cavity / Q000166:cytology; D010544:Peroxidases / Q000032:analysis*; D010984:Platinum
D015100:3,3'-Diaminobenzidine; D010984:Platinum; D010544:Peroxidases