Mechanism for antigen detection on deplasticized epoxy sections.
The purpose of this investigation was to explain why deplasticizing of epoxy sections gives higher immunogold labeling than non-deplasticizing. The methods used were the following: (1) Comparison of the ratio of immunogold labeling of deplasticized and non-deplasticized sections with gold particles of different sizes and comparison of this ratio with respect to sections of different thickness, (2) the tilt method (Brorson et al., 1994). Human kidney tissue with amyloid A depositions, human fibrin, and human pituitary tissue were embedded, sections were deplasticized on grids, treated with anti-Aa, anti-fibrinogen or anti-ACTH (ACTH = adrenocorticotropic hormone), and reembedded on grids. Indications of significant antibody penetration were found only at the periphery of structures (ACTH-vesicles). This penetration was about 30 nm. The ratios of immunogold labeling of deplasticized and non-deplasticized sections were approximately 2, 5 and 1 for amyloid, fibrin and ACTH, respectively, and were independent of the gold particle size. No significant differences of gold labeling were found between thicker and thinner deplasticized epoxy sections regardless the gold particle size. No significant differences of gold labeling between deplasticized epoxy sections and LR-White sections were found on interior areas of ACTH-vesicles or amyloid A plaques. The increased labeling of deplasticized epoxy sections compared to normal epoxy sections seemed to be mainly a surface phenomenon. The practical significance of this observation is that deplasticizing of epoxy sections may be a better method for localizing antigens at the periphery of structures than the use of other resin embedding media. Deplasticizing of epoxy sections may be a method of choice in a pathological laboratory to detect antigens in routinely embedded tissues.
Brorson
S H
SH
Department of Pathology, Ullevål Hospital, Oslo, Norway.
Skjørten
F
F
D000324:Adrenocorticotropic Hormone / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D000818:Animals; D000937:Antigen-Antibody Reactions; D000941:Antigens / Q000032:analysis*; D004853:Epoxy Resins / Q000737:chemistry; D000431:Ethanol / Q000031:analogs & derivatives* / Q000494:pharmacology; D006041:Goats; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism; D007150:Immunohistochemistry; D007668:Kidney / Q000737:chemistry / Q000648:ultrastructure; D051379:Mice; D016253:Microscopy, Immunoelectron / Q000379:methods; D008867:Microtomy; D010903:Pituitary Gland, Anterior / Q000737:chemistry / Q000648:ultrastructure; D011817:Rabbits; D000685:Serum Amyloid A Protein / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D012997:Solvents / Q000494:pharmacology*; D016610:Tissue Embedding / Q000379:methods*
D000941:Antigens; D004853:Epoxy Resins; D007074:Immunoglobulin G; D000685:Serum Amyloid A Protein; D012997:Solvents; C098088:sodium ethoxide; D000431:Ethanol; D000324:Adrenocorticotropic Hormone