Acetylation has been shown to be an important posttranslational modification (PTM) of both histone and nonhistone proteins with particular implications in cell signaling and transcriptional regulation of gene expression. Many studies have already demonstrated that SIRT1 is able to deacetylate histones and lead to gene silencing. It can also regulate the function of tumor suppressors including FOXO proteins and p53 by deacetylation. Here, we describe three experimental approaches for studying the modulation of the acetylation status of some of the known downstream targets of SIRT1.