All lanes : Anti-PKM2 (phospho Y105) antibody (ab156856) at 1 µg/mlLane 1 : F9 (Mouse embryonic carcinoma cell line) Whole Cell LysateLane 2 : F9 (Mouse embryonic carcinoma cell line) Whole Cell Lysate with Immunising peptide at 1 µg/mlLane 3 : F9 (Mouse embryonic carcinoma cell line) Whole Cell Lysate with Control peptide at 1 µg/mlLysates/proteins at 25 µg per lane.SecondaryGoat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilutiondeveloped using the ECL techniquePerformed under reducing conditions.
ab156856 was tested using an Indirect ELISA approach. The wells were coated with peptide (1µg/ml at 100µl/well) overnight at 4°C, followed by a 5% BSA blocking step for 1 hour at room temperature. The primary Ab was then added at a dilution range of 1- 0.00025µg/ml (100µl/well) for 1hr at room temperature. A HRP-conjugated anti-rabbit IgG (heavy and light chain) was used as a secondary antibody at 1:20,000 dilution for 1hr at room temperature.