Purpose The aim of this study was to investigate the wavefront aberration changes in human eyes caused by a gradient of increasing accommodation stimuli. Design This is a prospective, single-site study. Methods Healthy volunteers (n ¼ 22) aged 18-28 years whose refraction states were emmetropia or mild myopia, with astigmatism o1 diopter (D), were included in this study. After dilating the right pupil with 0.5% phenylephrine drops, the wavefront aberration of the right eye was measured continuously either without or with 1, 2, 3, 4, 5, or 6D accommodation stimuli (WFA1000B psychophysical aberrometer). The root mean square (RMS) values of the total wavefront aberrations, higher-order aberrations, and 35 individual Zernike aberrations under different accommodation stimuli were calculated and compared. Results The average induced accommodations using 1, 2, 3, 4, 5, or 6D accommodation stimuli were 0.848, 1.626, 2.375, 3.249, 4.181, or 5.085 D, respectively. The RMS of total wavefront aberrations, as well as higher-order aberrations, showed no significant effects with 1-3 D accommodation stimuli, but increased significantly under 4, 5, and 6 D accommodation stimuli compared with relaxed accommodation. Zernike coefficients of Z 0 4 significantly decreased with increasing levels of accommodation. Conclusion Higher-order wavefront aberrations in human eyes changed with increased accommodation. These results are consistent with Schachar's accommodation theory.