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Makeup is a prominent example of the universal human practice of personal decoration. Many studies have shown that makeup makes the face appear more beautiful, but the visual cues mediating this effect are not well understood. A widespread belief holds that makeup makes the facial features appear larger. We tested this hypothesis using a novel reference comparison paradigm, in which carefully controlled photographs of faces with and without makeup were compared with an average reference face. Participants compared the relative size of specific features (eyebrows, eyes, nose, mouth) of the reference face and individual faces with or without makeup. Across three studies we found consistent evidence that eyes and eyebrows appeared larger with makeup than without. In contrast, there was almost no evidence that the lips appeared larger with makeup than without. In two studies using professionally applied makeup the nose appeared smaller with makeup than without, but in a study using self-applied makeup there was no difference. Thus makeup was found to alter the facial feature sizes in ways that are related to age and sex, two known factors of beauty. These results provide further evidence to support the idea that makeup functions in part by modifying biologically based factors of beauty.