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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
GR Login, SJ Galli and AM Dvorak
Department of Pathology, Harvard School of Dental Medicine, Boston, Massachusetts.
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.
Volume 40,
Issue 9,
pp. 1247-1256,
09/01/1992
Copyright © 1992 by The Histochemical Society
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