Ultrastructural immunogold localization of tumor necrosis factor-alpha to the cytoplasmic granules of rat peritoneal mast cells with rapid microwave fixation

J Allergy Clin Immunol. 1994 Sep;94(3 Pt 1):531-6. doi: 10.1016/0091-6749(94)90210-0.


Tumor necrosis factor-alpha (TNF-alpha) is a multifunctional, proinflammatory cytokine, which can be produced by mast cells and several other cell types. We used a newly developed microwave energy-assisted aldehyde fixation method to prepare purified rat peritoneal mast cells for the postembedding immunogold ultrastructural localization of TNF-alpha. These fixation methods were superior to chemical fixation alone in preserving both the ultrastructural morphology and immunoreactive TNF-alpha in rat mast cells. The percent of TNF-alpha-positive mast cells in samples prepared with microwave-assisted fixation in low (84%) and standard (81%) glutaraldehyde concentrations exceeded that for low (56%) and standard (15%) glutaraldehyde concentrations without the assistance of microwave energy. TNF-alpha was identified in the large storage granules of rat mast cells. The percent of positive granules in microwave-assisted standard (44%) and low (40%) glutaraldehyde samples was considerably higher than the percent of positive granules in standard (5%) and low (10%) glutaraldehyde-fixed samples without microwave assistance. This location of TNF-alpha in rat peritoneal mast cells suggests that this cytokine can use the regulated secretory route(s) for release from appropriately stimulated rat mast cells into the microenvironment.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cytoplasmic Granules / chemistry*
  • Cytoplasmic Granules / ultrastructure
  • Immunohistochemistry*
  • Male
  • Mast Cells / chemistry*
  • Mast Cells / ultrastructure
  • Microscopy, Immunoelectron
  • Microwaves
  • Peritoneal Cavity / cytology
  • Rats
  • Rats, Sprague-Dawley
  • Tissue Fixation / methods*
  • Tumor Necrosis Factor-alpha / analysis*


  • Tumor Necrosis Factor-alpha