Fear overgeneralization and perceived uncertainty about future outcomes have been suggested as risk factors for clinical anxiety. However, little is known regarding how they influence each other. In this study, we investigated whether different levels of threat uncertainty influence fear generalization. Three groups of healthy participants underwent a differential fear conditioning protocol followed by a generalization test. All groups learned to associate one female face (conditioned stimulus, CS+) with a female scream (unconditioned stimulus, US) while the other face (CS-) was not associated with the scream. In order to manipulate threat uncertainty, one group (low uncertainty, n = 26) received 80%, the second group (moderate uncertainty, n = 32) received 60%, and the third group (high uncertainty, n = 30) 40% CS-US contingency. In the generalization test, all groups saw CS+ and CS- again as well as four morphs that varied in similarity with the CS+ in steps of 20%. Subjective (expectancy, valence, and arousal ratings), psychophysiological (skin conductance response, SCR), and visuocortical (steady-state visual evoked potentials, ssVEPs) indices of fear were registered. Participants expected the US in accordance with their reinforcement schedules but displayed stronger skin conductance with more uncertainty. However, acquisition of conditioned fear was not evident in ssVEPs. During the generalization test, we found no effect of threat uncertainty in any of the measured variables, but the strength of generalization for threat expectancy ratings was positively correlated with dispositional intolerance of uncertainty. This study suggests that mere threat uncertainty does not modulate fear generalization.