Shown below is an actin filament pelleting assay: actin purified from yeast was induced to polymerize with salt and magnesium in the presence (lanes 3-7) or absence (lanes 1 and 2) of Aip1p and in the presence of different amounts of cofilin. The filaments are then pelleted and the supernatant versus the pellet is sampled in a coomassie stained gel to quantify the amount of actin present in the filamentous form. As can bee seen in lane 1, actin alone efficiently forms filaments. The addition of cofilin at stoichiometric amounts (lane 2) induces a slight decrease in the amount of F-actin due to the cofilin induced increase in off rate from the minus ends of the filaments and a weak filament severing activity. Note the cofilin is largely present in the pellet fraction as it binds actin filaments 1:1 with the actin subunits. The addition of Aip1p at a 1:1 stoichiometry causes a large decrease in actin from the pellet phase and this effect is dependent on cofilin, little effect of Aip1p is observed once the cofilin is dropped down to 1/5 the amount of actin. This assay shows that Aip1p can act to destabilize actin filaments but only actin filaments that are optimally decorated with cofilin.