Immunohistochemical and biochemical characterization of antisera raised to human tumor nucleoli.
Antinucleolar antisera were raised in rabbits, goats and sheep to nucleoli isolated from three human tumor cell lines. The antisera were shown to cross-react by immunofluorescence with human tumor cell lines originating from different organs and with frozen sections from a wide variety of human malignant and non-malignant tissues. Tumor versus normal tissue discrimination by several antisera was significantly improved by treatment of frozen tissues with a buffered glutaraldehyde/Triton X-100 solution prior to immunofluorescent staining. The molecular specificity of these antisera was determined by immunostaining electrotransfer nitrocellulose strips following SDS-PAGE of nucleolar preparations and nuclear extracts. Although different immunostaining patterns were obtained for individual antinucleolar antisera, nucleolar proteins of molecular weight 120, 100, 94, 68, 54, 38, 33, and 32 kDa were the most often recognized by antisera raised in the three different species. G187 antiserum strongly reacted with 100, 94, and 38 kDa proteins from freshly obtained leukemic specimens. The Immunoreactivity of the 100, 94, and 38 kDa proteins was unaffected by glutaraldehyde/Triton X-100 treatment when immunostained with antisera that demonstrated the greatest tumor specificity on sections treated with glutaraldehyde/Triton X-100. These three nucleolar proteins may be more highly associated with nucleoli of malignant cells than with nucleoli of normal cells.
Greenfield
R S
RS
Bristol-Myers Company, Antitumor Immunology Department, Wallingford, CT 06492.
Lamon
M O
MO
Olech
L J
LJ
Grattan
J A
JA
D000818:Animals; D000951:Antigens, Neoplasm / Q000032:analysis*; D002466:Cell Nucleolus / Q000276:immunology*; D005455:Fluorescent Antibody Technique; D005976:Glutaral / Q000494:pharmacology; D006801:Humans; D007106:Immune Sera / Q000276:immunology*; D007150:Immunohistochemistry; D009687:Nuclear Proteins / Q000032:analysis; D011817:Rabbits; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D007106:Immune Sera; D009687:Nuclear Proteins; C040391:malignancy-associated nucleolar antigen, human; D005976:Glutaral