Team:Warsaw/Calendar-Main/8 September 2008

Our webpage has a completely new layout Emilia

Purification of proteins: A-alpha Piotr Electrophoresis in polyacrylamide gel (12 %) of sonicate containing A-alpha (Fig. 1.).

 Fig. 1. Sonicate containing A-alpha protein fusion The arrow shows place of our overexpressed protein: 1. Marker 2. sonicate A-alpha with induction

Michał L. 

Supernatant containing A-alpha was mixed with NiNta bead in proportion 10:1 and placed on rotating platform for 30 min at 4&deg;C Bead was spun down at 6000 RPM, 5 min at 4&deg;C Supernatant was discarded and equal amount of Elution buffer 1 (Sonication buffer A suplemented with 50 mM imidazole) was placed on top of the bead Bead was spun down at 6000 RPM, 5 min at 4&deg;C Supernatant was collected for further analysis and equal amount of Elution buffer 2 (Sonication buffer A supplemented with 100 mM imidazole) was placed on top of the bead The above two steps were repeated for elution buffers 3 and 4 (150 mM and 200 mM imidazole)</li> All fractions obtained in this procedure were resolved on 8% SDS-PAGE gel. It turned out that optimum imidazole concentration for elution of His-tagged A protein is 150 mM</li> <ol>