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The characterization and use of different antibodies against the hsp70 major heat shock protein family for the development of an immunoassay
Author(s) -
Margulis Boris A.,
Nacharov Peter V.,
Tsvetkova Olga I.,
Welsh Michael,
Kinev Alexander V.
Publication year - 1991
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150120913
Subject(s) - polyclonal antibodies , antiserum , immunoassay , hsp70 , monoclonal antibody , epitope , antibody , immunoprecipitation , microbiology and biotechnology , antigen , heat shock protein , affinity chromatography , chemistry , biology , biochemistry , enzyme , immunology , gene
The hsp70 family of major stress proteins is composed of several different members exhibiting similar structural and functional properties. In order to obtain an antiserum with wide epitope reactivity, rabbits were immunized with a mixture of native and denatured hsp70 purified from bovine muscle by ATP‐affinity chromatography. Screening for antibody specificity was performed by a “sandwich” enzyme linked immunosorbent assay (ELISA). Immunoprecipitation and immunoblotting analyses demonstrated that the polyclonal antiserum obtained by us and a monoclonal antibody raised against a different preparation of antigen recognized the same determinant on the native hsp70 molecule (inducible form). With a different specificity the polyclonal antiserum recognized only the denatured monomers of the other members of the hsp70 family. These results are discussed in relation to the immunological features of the hsp70 molecule and to the development of an immunoassay for the detection of hsp70 in Cell and tissue extracts.