Fig. 5

Use of anti-SIRT1 and anti-EZH2 antibodies in co-immunoprecipitation experiments. Anti-SIRT1 or anti-EZH2 antibody was added to total cell lysate as indicated, but omitted from samples labelled “IP -ve”. Immune complexes were captured on protein A/G Sepharose then eluted and analysed, along with an equivalent sample of the input protein, by Western blotting using anti-SIRT1 or anti-EZH2 antibody, as indicated. Both antibodies when used for immunoprecipitation and Western blotting confirmed self-enrichment of the corresponding protein in the immunoprecipitated samples but neither led to enrichment of the other protein. Approximate molecular weights of proteins seen as specific bands are indicated. Data using anti-SIRT1 antibody are for Caco-2 cells and are consistent with data obtained using HuVECs. Data using anti-EZH2 antibody are for HuVECs and are consistent with data obtained using Caco-2 cells