Monoclonal antibodies against distinct determinants of histone H5 bind to chromatin.
A series of monoclonal antibodies specific for distinguishable epitopes in chromosomal protein histone H5 were obtained from mice immunized with either free H5 or H5 . RNA complexes. The antibodies elicited by H5 could be distinguished from those elicited by H5 . RNA by their binding to native or acid-denatured H5, by their interaction with the globular region of H5, and by their cross-reactivity with H1o. The specificity of the antibodies was assessed by enzyme-linked immunosorbent assay (ELISA) and immunoblotting experiments. The antibodies could distinguish between H5 and the closely related histones H1 and H1o. The binding of some of the antibodies to the antigens was dependent on the type of assay used, suggesting nonrandom binding of the antigen to the solid supports used in ELISA and immunoblotting. Competitive ELISA experiments indicate that 8 of the 11 antibodies characterized bind to distinct epitopes. Three monoclonal antibodies bind to epitopes which are in close spatial proximity, causing mutual steric hindrance. The monoclonal antibodies bind to nuclei of fixed cells and to isolated chromatin, indicating that the epitopes are present both in the purified protein and in chromatin-complexed H5. These monoclonal antibodies can be used to study the organization of distinct regions of histones H5 and H1o in chromatin and chromosomes.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D002843:Chromatin / Q000032:analysis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D005260:Female; D005455:Fluorescent Antibody Technique; D006657:Histones / Q000032:analysis* / Q000276:immunology; D005779:Immunodiffusion; D051379:Mice; D008807:Mice, Inbred BALB C
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D002843:Chromatin; D000939:Epitopes; D006657:Histones