Detecting peroxiredoxin hyperoxidation by one-dimensional isoelectric focusing

The activity of typical 2-cys peroxiredoxin (Prxs) can be regulated by hyperoxidation with a consequent loss of redox activity. Here we developed a simple assay to monitor the level of hyperoxidation of different typical 2-cys prxs simultaneously. This assay only requires standard equipment and can compare different samples on the same gel. It requires much less time than conventional 2D gels and gives more information than Western blotting with an antibody specific for hyperoxidized peroxiredoxin. This method could also be used to monitor protein modification with a charge difference such as phosphorylation.