Itch and pain are both mediated by small sensory fibres. Atopic dermatitis (AD) patients usually report stress-induced flares, but the impact of stress on sensory fibres in lesional and non-lesional skin remains inconclusive. This observational study assessed the effect of acute stress on sensory profiles in subjects with AD (n = 18) and healthy controls (HC, n = 21). Participants completed clinical and psychological questionnaires, and quantitative sensory testing was performed on lesional and non-lesional skin in AD and healthy skin in HC. Assessments were done before and after the Montreal Imaging Stress Task, an acute stress protocol. Stress responses were evaluated by anxiety ratings, heart rate (HR) and salivary cortisol (CORT). Cortisol binding globulin (CBG) was quantified as an indirect measure for circulating CORT. AD participants reported higher anxiety, depression and stress perception than HC. HR was similar between groups, but AD participants showed a blunted CORT response post-stress and lower CBG levels, suggesting altered stress regulation. Acute stress reduced cold sensitivity in HC and non-lesional AD skin but had no effect on lesions. These findings indicate that the effects of stress on small fibres depend on the condition of the skin and emphasise the sensory alterations experienced by AD patients.