In this study we investigate the effect of programmed cross-flows on the error in the hydrodynamic radii (rh) determination with asymmetrical flow field-flow fractionation (AsFlFFF). Three different standard polystyrene particles (nominal radii of 30 and 40 and 50 nm) are fractionated with exponentially and linearly decaying cross-flows with different decay rates. Hydrodynamic radii are calculated according to retention theory including steric effects. Rapid decay is expected to give rise to systematic deviations in rh determination. The error in rh was found to be small when decay rates with half-lives longer than 6 min were used, whereas steeper decays could give rise to errors as high as 16% of the particle size. The error is often explained in terms of secondary relaxation. However, comparisons show that experimental errors are significantly larger than what would be expected due to secondary relaxation, suggesting that other factors also have to be considered in order to fully understand deviations for rapidly decaying cross-flow.