A Simplified Method to Assay Protein Carbonylation by Spectrophotometry

Protein carbonylation is an irreversible oxidation process leading to a loss of function of carbonylated proteins. Carbonylation is largely considered as a hallmark of oxidative stress, the level of protein carbonylation being an indicator of the oxidative cellular status. The method described herein represents an adaptation to the commonly used 2,4-dinitrophenylhydrazine (DNPH)-based spectrophotometric method to monitor protein carbonylation level. The classical final sample precipitation was replaced by a gel filtration step avoiding the tedious and repetitive washings of the protein pellet to remove free DNPH while allowing optimal protein recovery.This improved protocol here implemented to assay protein carbonylation in plant leaves can potentially be used with any cellular extract.