All lanes : Anti-Geminin antibody ( ab12147 ) at 1/500 dilution Lane 1 : 293 lysate Lane 2 : 293 lysate with Human Geminin peptide (ab12408) at 1 µg/ml Lysates/proteins at 5 µg/ml per lane. developed using the ECL technique Performed under reducing conditions. Predicted band size : 23.5 kDa Observed band size : 33 kDa ( why is the actual band size different from the predicted? ) The lower band represents Geminin (33 kDa - predicted molecular weight 23 kDa), and is blocked by the immunising peptide, the upper band is likely to be non-specific as it is not blocked by the peptide.