Improved immunogold labeling of epoxy sections by the use of propylene oxide as additional agent in dehydration, infiltration and embedding.
The purpose of this study was to examine how the intensity of the immunogold labeling on epoxy sections was affected by the use of propylene oxide as an agent in addition to ethanol in the dehydration and infiltration, and also to examine the effect on the immunogold labeling by adding small amounts of propylene oxide to the embedding mixture. Increased knowledge of the mechanism for antigen detection on resin sections was another aim. Thyroid tissue, kidney tissue, and fibrin were embedded in epoxy resin; some with ethanol as the only dehydration agent and others with propylene oxide as an additional agent in dehydration, infiltration or embedding steps in different ways. Immunogold labeling was performed with anti-thyroglobulin, anti-IgG, and anti-fibrinogen, respectively. A higher degree of immunogold labeling was achieved by increasing the concentration of accelerator during infiltration and embedding (Brorson and Skjørten, 1996a, Micron, 27, 211-217). The immunogold labeling of the sections that were based on additional dehydration and infiltration with propylene oxide showed significantly more intense labeling than the sections of tissues that had only been exposed to ethanol in the dehydration and infiltration steps. The embedding of tissues in a mixture of epoxy resin and 5-10% propylene oxide gave higher yields of immunogold labeling than if pure epoxy resin was used for the embedding. The improved labeling is explained by higher amplitudes of protruding antigens on the surface of the sections because antigens are less tightly incorporated in the polymer network when using propylene oxide as additional agent in dehydration, infiltration or embedding. These results illustrate the advantage of using propylene oxide as an additional agent when preparing specimens for immunoelectron microscopy with epoxy resin embedding.
Brorson
S H
SH
Department of Pathology, Ullevål Hospital, Oslo, Norway.
D000818:Animals; D004852:Epoxy Compounds / Q000378:metabolism*; D005337:Fibrin / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D007668:Kidney / Q000648:ultrastructure; D008223:Lymphoma / Q000473:pathology; D016253:Microscopy, Immunoelectron / Q000379:methods; D013552:Swine; D013961:Thyroid Gland / Q000473:pathology / Q000648:ultrastructure; D016610:Tissue Embedding / Q000379:methods*; D016707:Tissue Fixation / Q000379:methods*
D004852:Epoxy Compounds; D005337:Fibrin; C009068:propylene oxide