Visual discomfort and fatigue due to accommodation-convergence (AC) conflict in stereoscopic displays has been widely reported, but little is known about its impact on visual acuity, particularly when automotive three-dimensional (3D) head-up displays (HUDs) are involved. This paper presents a study on the visual acuity response when an indigenously developed 75% transparent retroreflective screen is used as a windshield 3D HUD. The simulated optical collimation technique was used to provide the virtual content at a farther depth (i.e. on the road while driving). Two user test experiments were performed. The first test was performed under the see-through condition, where the real scene (i.e. roadside view) was perceived through the 3D HUD, while the second test was performed under the simulated collimation condition, where a stereo-collimated virtual content was projected on the HUD at a farther depth. The results showed a slightly declining trend (from 20/20 to 20/25) in visual acuity response when the HUD screen was placed between the viewer and the scene. An inverse relation between the amount of AC conflict and visual acuity was observed under the simulated collimation condition. The > 100 cm user-to-screen distance was found to be comfortable, providing the highest acuity response.