Gas6 and the Receptor Tyrosine Kinase Axl in Clear Cell Renal Cell Carcinoma (original) (raw)
Figure 1
Axl is phosphorylated and Gas6 is γ-carboxylated in human ccRCC tissue.
Homogenized tissue lysates from a panel (patient one to eight) of matched ccRCC tissues and unaffected kidney cortex counterparts were used for Axl (A) and Gas6 (B) immunoprecipitation with specific polyclonal antisera (#R042 towards Axl N-terminal; #005 towards Gas6) containing 50–100 µg total immunoglobulin molecules (about 0.5–1.0 µg specific immunoprecipitating antibody). Pellets were separated on a 6% denaturing SDS-PAGE gel. Protein expression and posttranslational modifications were verified with western blot analysis using Axl (Axl-C20; towards Axl C-terminal), Gas6 (R&D), phosphotyrosine (PY99) and Gla (M3b; 5 mM EDTA final) specific antibodies, respectively. Membranes were first immunoblotted for phosphotyrosine and Gla, respectively, and thereafter stripped and reprobed for total Axl and Gas6. For each patient the matched samples are denoted: N (normal unaffected kidney cortex) and T (ccRCC tissue). Absolute concentrations in the lysates measured by ELISA are depicted below each sample.