Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with special importance in reproductive and developmental processes. PACAP is found in two bioactive forms: PACAP27 and PACAP38. Recently, we have described that PACAP38 is present in high levels in the milk of human and ruminant animals. Breastfeeding is of utmost importance in proper nutrition of the newborn, but artificial nursing with infant formulas is necessary when breastfeeding is not available. Composition of the breast milk varies during the whole period of nursing and it shows differences at the beginning (foremilk) and the end of an actual suckling (hindmilk). The aim of this study was to investigate PACAP38-like immunoreactivity (PACAP38-LI) in different milk and infant formula samples by radioimmunoassay and to prove the presence of PACAP38 in the infant formula by mass spectrometry. We found similar PACAP38-LI in human mature foremilk and hindmilk samples, in the fresh and pasteurized cow milk and also in formulas. However, we found significantly higher PACAP38-LI in the hypoantigenic formula undergoing extensive hydrolysis compared to the non-hypoantigenic ones. Our results suggest that PACAP38 is relatively stable in the milk and it can withstand the manufacturing processes.