Building-related symptoms (BRS) is a significant work-related and public health problem, characterized by non-specific symptoms occurring in a particular building. The cause of BRS is unknown, but certain reactive compounds are suggested risk factors. The aim of this controlled exposure study was to investigate whether BRS cases report more odor annoyance and symptoms and show altered autonomous nervous system (ANS) response during exposure to the reactive aldehyde, acrolein in comparison with referents. Individuals with BRS (n = 18) and referents (n = 14) took part in two exposure sessions (80 min). One session contained heptane alone, and the other heptane and acrolein. Perceived odor annoyance; eye, nose, and throat symptoms; and ANS response were measured continuously. BRS cases did not experience more odor annoyance; eye, nose, and throat symptoms; or altered ANS response in comparison with referents during the exposures. Supplementary analyses revealed that BRS cases that also reported chemical intolerance perceived more symptoms than referents during acrolein exposure. Acrolein exposure at a concentration below previously reported sensory irritation detection thresholds is perceived as more irritating by a subgroup of BRS individuals compared with referents. The results of this study indicate that a subset of individuals with building related symptoms (BRS) has a lowered sensory irritation threshold towards acrolein exposure. Future guidelines on chemical exposures to acrolein should take time and individual sensitivity into account.