Selection of a fixative for identifying T cell subsets, B cells, and macrophages in paraffin-embedded mouse spleen.
Fixation techniques were investigated for their ability to preserve morphology, esterase activity and cell surface antigens in paraffin-embedded mouse lymphoid tissue. The avidin-biotin-peroxidase system was used to stain antigens Thy-1.2, Lyt-1, Lyt-2, RA32C2 and F4/80. Conventional fixatives were compared with fixatives containing periodate and lysine plus paraformaldehyde and/or glutaraldehyde. Conventional fixatives preserved esterase activity but not many cell surface antigens. Periodate-lysine fixatives allowed better preservation of membrane antigens, but esterase activity was often lost at antigen-preserving concentrations of paraformaldehyde or glutaraldehyde. However, a periodate-lysine fixative containing both paraformaldehyde and glutaraldehyde preserved esterase and showed good to excellent staining of Lyt-1, Thy-1.2, RA32C2, and F4/80. Lyt-2 could not be stained with any fixative, but was well preserved in frozen material post-fixed with periodate-lysine based fixatives. We conclude that with proper fixation immune cell surface markers can be identified in paraffin-embedded tissue.
D000818:Animals; D001402:B-Lymphocytes / Q000145:classification / Q000201:enzymology / Q000276:immunology*; D005404:Fixatives; D006651:Histocytochemistry; D007124:Immunoenzyme Techniques; D008239:Lysine; D008264:Macrophages / Q000145:classification / Q000201:enzymology / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D010232:Paraffin; D010504:Periodic Acid; D013154:Spleen / Q000166:cytology; D013601:T-Lymphocytes / Q000145:classification / Q000201:enzymology / Q000276:immunology*
D005404:Fixatives; D010504:Periodic Acid; D010232:Paraffin; C009288:metaperiodate; D008239:Lysine