(A and B) Representative Western blot assays performed using anti Flag monoclonal antibody, the pNCC-T58 phosphoantibody, and β-actin antibody, as stated. Total proteins were extracted from Xenopus laevis oocytes injected with water or rNCC cRNA and exposed to insulin for 0, 5, 15 and 30 minutes (A), or to 15 minutes in the absence or presence of wortmannin (B) (C) A representative Western blot analysis of protein extracts from ex-vivo perfused rat-kidneys. The kidneys were perfused with isotonic control solution or with isotonic solution containing 0.250 U/mL of insulin. (D) Densitometry analysis of phosphor-NCC over total rNCC from three different Western blot analyses compiling a total of 10 controls and 16 insulin perfused kidneys. (*p<0.01 vs control)