We describe the use of the fast Fourier transform (FFT) in the measurement of anisotropy in electrospun scaffolds of gelatin as a function of the starting conditions. In electrospinning, fiber alignment and overall scaffold anisotropy can be manipulated by controlling the motion of the collecting mandrel with respect to the source electrospinning solution. By using FFT to assign relative alignment values to an electrospun matrix it is possible to systematically evaluate how different processing variables impact the structure and material properties of a scaffold. Gelatin was suspended at varying concentrations (80, 100, 130, 150mg/ml) and electrospun from 2,2,2 trifluoroethanol onto rotating mandrels (200-7000 RPM). At each starting concentration, fiber diameter remained constant over a wide range of mandrel RPM. Scaffold anisotropy developed as a function of fiber diameter and mandrel RPM. The induction of varying degrees of anisotropy imparted distinctive material properties to the electrospun scaffolds. The FFT is a rapid method for evaluating fiber alignment in tissue-engineering materials. (c) 2006 Elsevier Ltd. All rights reserved.