In colloidal suspensions containing a binary mixture of hard spheres depletion forces occur which substantially contribute to the interaction of the larger spheres among themselves and a wall, respectively. We investigated the depletion force acting on a large colloidal polystyrene sphere immersed in a solution of small, noncharged polymer coils close to a flat glass surface by means of total internal reflection microscopy. When the distance between the polystyrene sphere and the wall is smaller than the diameter of the polymer coils, an attractive potential acting on the sphere is observed which depends strongly on the polymer concentration. Our results are in agreement with theoretical predictions.[S0031-9007 (98) The stability of colloidal mixtures consisting of larger and smaller particles is well known to be strongly influenced by entropic depletion forces. Accordingly, those forces are essential in understanding phase separation phenomena and flocculation of binary hard-sphere mixtures, colloids in the presence of micelles, and of colloid polymer mixtures [1,2]. Very recently, it was suggested that depletion forces even may play an important role in the shape changes of phospholipid vesicles [3].The principal phenomenon of depletion interaction is easily understood when, e.g., a hard sphere of radius R suspended in a fluid containing smaller spheres of radius r (the latter are referred to as macromolecules in the following) in front of a wall at distance z is considered (see Fig. 1). If z decreases below the diameter of the macromolecules they are expelled from the region between the sphere and the wall. Consequently, the concentration of macromolecules becomes depleted in this region compared to that of the bulk, and an effective osmotic pressure causing a net attraction between the sphere and the wall occurs. Such an attractive force is also observed when the wall is replaced by another hard sphere.The first quantitative explanation of this effect was given by Asakura and Oosawa [4]. According to their calculation the change in the Helmholtz free energy DF of a single sphere positioned at distance z from a wall and suspended in a fluid of macromolecules can be written as