The fatty acids (FAs) of human milk (HM) are the building blocks of the HM lipidome, contributing to infant health and development; however, this has not been comprehensively charac-terised with respect to infant intake. Eighteen Western Australian mother–infant dyads provided monthly longitudinal HM samples during six months of exclusive breastfeeding. Monthly anthro-pometric measurements, health data and basic maternal food frequency data were also collected. At three months, infant 24 h milk intake and total lipid intake were measured. The FA profile was analysed using gas chromatography–mass spectrometry. Linear regression and Pearson’s correlation were used to identify associations between HM FA composition, HM FA intake, maternal characteristics and infant growth and developmental outcomes. Mean infant intake of total lipids was 29.7 ± 9.4 g/day. HM FA composition exhibited wide variation between dyads and throughout lactation. Infant intake of a number of FAs, including C15:0, C18:1, C18:2 and C20:3, was positively related to infant growth (all p < 0.001). There were no relationships detected between C22:5 and C20:5 and infant head circumference. Infant total lipid intake and the infant intake of many FAs play essential roles in infant growth and development. This study highlights the important relationships of many HM FAs not previously described, including C15:0 and C18:2 species. Infant outcomes should be considered in the context of intake in future HM studies.