PMID: 6166621Jun 1, 1981Paper

Use of colloidal gold particles in double-labeling immunoelectron microscopy of ultrathin frozen tissue sections

The Journal of Cell Biology
H J GeuzeR C Scheffer

Abstract

Complexes of protein-A with 5 and 16 nm colloidal gold particles (PA/Au5 and PA/Au16) are presented as sensitive and clean immunoprobes for ultrathin frozen sections of slightly fixed tissue. The probes are suitable for indirect labeling and offer the opportunity to mark multiple sites. The best procedure for double labeling was to use the smaller probe first, i.e., antibody 1 - PA/Au5 - antibody 2 - PA/Au16. When this was done, no significant interference between PA/Au5 and PA/Au16 occurred. Using this double-labeling procedure we made an accurate comparison between the subcellular distributions of amylase as a typical secretory protein and of GP-2 a glycoprotein, characteristic for zymogen granule membrane (ZGM) preparations. We prepared two rabbit antibodies against GP-2. One antibody (R x ZGM) was obtained by immunizing with native membrane material. The specificity of R x ZGM was achieved by adsorption with the zymogen granule content subfraction. The other, R x GP-2, was raised against the GP-2 band of the SDS polyacrylamide profile of ZGM. We found that the carbohydrate moiety of GP-2 was involved in the antigenic determinant for R x ZGM, while R x GP-2 was most likely directed against GP-2 polypeptide backbone. THe immu...Continue Reading

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Citations

Jun 1, 1991·Journal of Electron Microscopy Technique·R J JiaoZ H Zhai
Jun 1, 1991·Journal of Electron Microscopy Technique·C HartfieldK Fischer Lindahl
Apr 1, 1990·Cell and Tissue Research·D Daga GordiniG M Bressan
Jan 1, 1984·Archives of Dermatological Research·D SchmittD Tuffery
Jan 1, 1986·Histochemistry·H Galjaard
Mar 1, 1983·The Histochemical Journal·F T Bosman
Sep 1, 1982·The Histochemical Journal·G M HodgesL K Trejdosiewicz
Jan 1, 1992·The Histochemical Journal·E K KarhukorpiK Väänänen
Jan 1, 1985·Journal of Inherited Metabolic Disease·H J SipsE E Sterchi
Oct 15, 1986·Experientia·H Galjaard
Jan 1, 1988·Journal of Steroid Biochemistry·F NaftolinL M Garcia-Segura
Jan 1, 1988·Journal of Virological Methods·A D HyattJ R White
Jan 1, 1996·Journal of Virological Methods·J M MackenzieP R Young
Jan 1, 1990·Electron Microscopy Reviews·I RaskaL Salamin-Michel
Oct 20, 2007·Nature Protocols·Jan W Slot, Hans J Geuze
Apr 7, 2009·Proceedings of the National Academy of Sciences of the United States of America·Nadine van MontfoortFerry Ossendorp

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