Fig 2. Protein radical detection in PSII membranes deprived of Mn₄O₅Ca complex using immuno-spin trapping technique.

Blot images show effects of [A] DPC and desferal, and [B] high pH on P^• formation. Size of the marker is indicated on the left side and the identification of bands, which has been done on the same gel, is presented on the right side. For immuno-spin trapping detection of DMPO-protein nitrone adducts, anti DMPO-protein nitrone adduct antibody was used and for the identification of bands, respective antibodies were used. PSII membranes deprived of Mn₄O₅Ca complex (10 μg Chl) were exposed to high light (1000 μmol photons m^-2 s^-1) in presence of 50 mM DMPO and 40 mM MES buffer (pH 6.5) for 30 min. In some experiments, 500 μM DPC or 50 μM desferal were added to the sample before illumination. For high pH effect, PSII membranes deprived of Mn₄O₅Ca complex were exposed to high light in buffers of different pH (40 mM HEPES buffer pH 7.5, 40 mM HEPES buffer 8.5 and40 mM CAPSO buffer pH 9.5). These blots are the representative images of three experiments.