Immunocytochemical localization of histamine in secretory granules of rat peritoneal mast cells with conventional or rapid microwave fixation and an ultrastructural post-embedding immunogold technique.
We used a post-embedding immunogold labeling approach to define the fine-structural localization of histamine in rat peritoneal mast cells that were fixed using either standard aldehyde fixation or a fast microwave-aldehyde fixation method. Specimens were processed routinely for electron microscopy. Thin sections were exposed first to guinea pig antihistamine antiserum and then to gold-conjugated goat IgG directed against guinea pig IgG. By transmission electron microscopy, gold particles were localized to the matrix of cytoplasmic granules. Control sections treated with non-immune sera did not show labeling of mast cells. Adsorption of antihistamine antiserum with purified histamine or histamine bound to agarose showed a significant reduction (p less than 0.005) in granule staining. We also confirmed that our isolation procedures yielded functionally competent mast cells which released histamine when stimulated with sheep anti-rat IgE antiserum or with compound 48/80. These studies define the conditions of fixation for electron microscopy that are appropriate for the localization of histamine in the granule matrix of rat peritoneal mast cells.
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G R
GR
Department of Pathology, Harvard School of Dental Medicine, Boston, Massachusetts.
Galli
S J
SJ
Dvorak
A M
AM
D000818:Animals; D003594:Cytoplasmic Granules / Q000378:metabolism* / Q000648:ultrastructure; D006046:Gold; D006632:Histamine / Q000378:metabolism*; D007150:Immunohistochemistry; D008297:Male; D008407:Mast Cells / Q000648:ultrastructure*; D008854:Microscopy, Electron; D008872:Microwaves; D010529:Peritoneal Cavity / Q000166:cytology; D051381:Rats; D016707:Tissue Fixation
D006046:Gold; D006632:Histamine