Figure 1: Treatment of NIH/3T3 cells with PDGF #9909 stimulates phosphorylation of Akt2 at Ser474, detected by PathScan ® Phospho-Akt2 (Ser474) Sandwich ELISA Kit (Mouse Preferred) #7932, but does not affect levels of total Akt2 protein detected by PathScan ® Total Akt2 Sandwich ELISA kit (Mouse Preferred) #7930. The absorbance readings at 450 nm are shown in the top figure, while the corresponding western blots using Akt2 (5B5) Rabbit mAb #2964 (left panel) and Phospho-Akt (Ser473) (193H12) Rabbit mAb #4058 (right panel) are shown in the bottom figure.
Figure 2: Demonstration of phospho-Akt protein sandwich ELISA kit specificity using recombinant phosphorylated Akt1, Akt2 and Akt3 proteins. Phospho-Akt1 (Ser473) is detected by #7160 while #7932 measures levels of phospho-Akt2 (Ser474). Recombinant phosphorylated Akt protein (1.0 ng per microwell) is assayed using both ELISA kits.
Figure 3: The relationship between protein concentration of lysates from untreated or PDGF-treated NIH/3T3 cells and the absorbance at 450 nm is shown. After starvation, NIH/3T3 cells (85% confluence) were treated with PDGF #9909 (50 ng/ml) for 10 min at 37ºC and then lysed.