A monoclonal antibody against insect CALNUC recognizes the prooncoprotein EWS specifically in mammalian cells. Immunohistochemical and biochemical studies of the antigen in rat tissues

Citation
J. Kawano et al., A monoclonal antibody against insect CALNUC recognizes the prooncoprotein EWS specifically in mammalian cells. Immunohistochemical and biochemical studies of the antigen in rat tissues, HISTOCHEM C, 115(5), 2001, pp. 421-428
Citations number
17
Categorie Soggetti
Medical Research Diagnosis & Treatment
Journal title
HISTOCHEMISTRY AND CELL BIOLOGY
ISSN journal
09486143 → ACNP
Volume
115
Issue
5
Year of publication
2001
Pages
421 - 428
Database
ISI
SICI code
0948-6143(200105)115:5<421:AMAAIC>2.0.ZU;2-V
Abstract
A monoclonal antibody against insect CALNUC was shown to recognize an 85-kD a nuclear protein specifically in mammalian cells. Amino acid sequencing of the protein purified from rat liver revealed it to be EWS, a prooncoprotei n for Ewing sarcomas and related tumors. Using the antibody, distribution o f EWS was studied in rat tissues fixed with 4% paraformaldehyde by immunohi stochemical methods. On thaw-fixed cryosections or those of perfusion-fixed tissues, almost all cell nuclei showed the specific staining. In immersion -fixed tissues, the staining unexpectedly disappeared in particular tissues (kidney cortex, liver, etc.), although it was recovered by autoclaving the cryosections. Western blotting also demonstrated the ubiquitous expression of EWS in the tissues. In extracts from the liver, the 85-kDa band rapidly disappeared in a Ca2+-dependent manner, but never in the testis. The antig en was very labile in kidney homogenates even without Ca2+. Biochemical stu dies with digoxigenin-labeled EWS showed that the Ca2+-dependent disappeara nce was associated with upward mobility shifts of EWS. These suggested that EWS was ubiquitously expressed in rat tissues, and that the antigen was ma sked in particular tissues during the immersion fixation.