Determination of enzymatic activity by zymogram and immunoblotting. Supernatant samples from P. pastoris SMD1168H transformants pGAPZαA (background expression) and pGAPZαA-SOD were collected after incubation for 96 hours at 28.5°C. As a positive control, 3 U of SOD were used. A. SOD activity under non-denaturing conditions. Concentrated protein samples (6×) were separated in native-PAGE 12%. SOD activity is visualized as white bands (asterisk). B. Protein samples (1×) were analyzed by immunoblotting using myc-specific (Myc) antisera.