Evaluating eIF4E/eIF4A Complexes by Co-IP

eIF4E/eIF4A complexes were evaluated by co-immunoprecipitation assays under different treatments. For these immuno-precipitation assays, cellular fractions (300 µg) were incubated with the antibody anti-eIF4A (1:400) for 2 h at 4 °C. Then, immune complexes were precipitated with protein G Agarose Fast Flow (Millipore) 12 h at 4 °C, according to the previous protocol [36 (link)]. Immuno-precipitated proteins were washed 3 times, and suspended in Laemmli buffer, separated by SDS-PAGE gels and transferred to PVDF membranes for WB analysis. eIF4E detection was performed based on a previously described protocol.