We measure all nonzero elements of the three-dimensional (3D) diffusion tensor D for clusters of colloidal spheres to a precision of 1% or better using digital holographic microscopy. We study both dimers and triangular trimers of spheres, for which no analytical calculations of the diffusion tensor exist. We observe anisotropic rotational and translational diffusion arising from the asymmetries of the clusters. In the case of the three-particle triangular cluster, we also detect a small but statistically significant difference in the rotational diffusion about the two in-plane axes. We attribute this difference to weak breaking of threefold rotational symmetry due to a small amount of particle polydispersity. Our experimental measurements agree well with numerical calculations and show how diffusion constants can be measured under conditions relevant to colloidal self-assembly, where theoretical and even numerical prediction is difficult.Diffusion plays a critical role in the dynamics, selfassembly, and rheology of complex fluids. In systems such as colloidal suspensions, which typically have shortranged interaction potentials, diffusion can in fact play a larger role than energy barriers in setting transition rates [1]. However, the diffusion of geometrically anisotropic particles, a common class of colloidal suspension that can also arise as intermediates in the selfassembly of spherical particles, can be difficult to predict. Theoretically determining friction factors for these particles requires analytically solving Stokes' equation, which is only possible for highly symmetric particles such as ellipsoids [2] or sphere dimers [3] in unbounded fluids. Numerical methods such as bead modeling [4] or finiteelement methods [5][6][7] require approximating the shape of the particles or the hydrodynamic interactions. These methods are difficult to apply when asymmetric particles diffuse near rigid boundaries or other particles, two situations that are relevant to colloidal self-assembly and dynamics in general. Thus experimental measurements of diffusion tensors are crucial.In particular, precision measurements on single particles rather than ensembles are necessary. Anisotropic particles show multiple diffusion timescales that are difficult to resolve through bulk techniques such as depolarized dynamic light scattering [8]. But there have been few single-particle studies of anisotropic diffusion in 3D. Video microscopy has been used to measure twodimensional (2D) diffusion of colloidal ellipsoids [9,10] and planar sphere clusters [11] but the technique yields limited information about out-of-plane motions [12][13][14]. Confocal microscopy can be used to study the 3D dynamics of geometrically anisotropic particles, but has only * vnm@seas.harvard.edu been applied to highly symmetric particles [15,16] and is limited by the time (∼ 1 s) needed to acquire a 3D stack. This can make it challenging to probe timescales comparable to particle diffusion times or to study rare processes such as the early stages of self-assem...