Fig. 5

Western blots analysis of hGR α , hGR β , and hsp90 performed using total protein extracted from HeLa cells after pre-adsorption with nonimmune rabbit IgG and anti-hGR α and -hGR β antibodies, and precipitation with protein A-Sepharose

Aliquots (100 µl) were incubated with nonimmune rabbit IgG and anti-hGRα-, and -hGRβ-specific antibodies and precipitated with protein A-Sepharose, as described in Materials and Methods and shown at the top. After washing, the immunoadsorbed/precipitated receptors were solubilized in SDS-sample buffer, each sample was divided into aliquots and resolved by SDS-PAGE. After transfer to nitrocellulose membranes, samples were immunoblotted with anti-hGRα (left panel), -hGRβ (middle panel) and -hsp90 (right panel). Anti-hGRβ antibody precipitated hGRα and hsp90, while anti-hGRα antibody precipitated hGRβ and hsp90. The position of identified immunoreactive bands are indicated against molecular weights standards.