Establishment and characterization of monoclonal antibody against androgen receptor.
Hybrid cell lines were prepared by the fusion of BALB/c myeloma NS-1 cells with the lymphocytes of BALB/c mice that were immunized with partially purified androgen receptor (AR) from human prostates. Nine clones of the hybrid progeny were determined for the production of antibodies against AR by immunoprecipitation assay. One of the clones, referred to as "5F4", was chosen for analysis of the detailed specificity. The clone "5F4" secreted IgM class antibodies against AR. Competition study demonstrated that "5F4" antibody inhibited androgen binding of AR, suggesting that the antibody identifies androgen binding site of AR. Immunoblotting analysis showed that the antibody identified the ARs as two proteins, 95 kD and 41 kD proteins, on a sodium dodecyl sulfate polyacrylamide gel. It is suspected that a 95 kD protein should be a monomeric AR and a 41 kD protein is a proteolytic fragment of AR. Immunohistochemical analysis demonstrated that androgen-dependent tissues--human prostatic hypertrophy tissues, an AR abundant prostatic cancer tissue and fibroblast cells from human genital skin--were stained intensely with "5F4" monoclonal antibody, while androgen-independent tissues--fibroblast cells from lymph nodes, an AR deficient prostatic cancer tissue and human prostatic cancer cell line, PC-3--showed no staining. These results also support the specificity of the antibody for AR.
Demura
T
T
Department of Urology, Hokkaido University School of Medicine, Sapporo, Japan.
Kuzumaki
N
N
Oda
A
A
Fujita
H
H
Ishibashi
T
T
Koyanagi
T
T
D000728:Androgens / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D000937:Antigen-Antibody Reactions; D001667:Binding, Competitive; D015153:Blotting, Western; D004306:Dose-Response Relationship, Immunologic; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007124:Immunoenzyme Techniques; D008297:Male; D011467:Prostate / Q000032:analysis / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000378:metabolism; D011944:Receptors, Androgen / Q000276:immunology* / Q000378:metabolism
D000728:Androgens; D000911:Antibodies, Monoclonal; D011944:Receptors, Androgen