Linker histones from grapevine were purified by electrophoretic methods (SDS-PAGE and electrotransfer onto a nitrocellulose sheet). Individual linker histones were recovered after nitrocellulose solubilization by acetone. The proteins precipitated after this treatment were used as antigen for subsequent immunizations of mice. Such purified immunogens were injected into mice with dimethyl sulfoxide, as the presence of nitrocellulose-protein complexes in the antigen pellets after the acetone treatment was suspected. The resulting antisera were specific to the injected antigens after only the first immunization and could be used as specific probes in immunohistological studies. Our approach seems more efficient (in using less antigen and obtaining a faster response) than the classical procedure recommended for histones in general (B. D. Stollar and M. Ward, 1970, J. Biol. Chem. 245, 1261-1266) and other methods that use free linker histone as immunogen.
Copyright 1998 Academic Press.