Background & Methods To examine the relationship between breastfeeding and maternally-rated infant temperament at age 3 months, 316 infants in the prospective Cambridge Baby Growth Study, UK had infant temperament assessed at age 3 months by mothers using the Revised Infant Behavior Questionnaire, which produces scores for three main dimensions of temperament derived from 14 subscales. Infant temperament scores were related to mode of infant milk feeding at age 3 months (breast only; formula milk only; or mixed) with adjustment for infant's age at assessment and an index of deprivation. Results Infant temperament dimension scores differed across the three infant feeding groups, but appeared to be comparable between exclusive breast-fed and mixed-fed infants. Compared to formula milk-fed infants, exclusive breast-fed and mixed-fed infants were rated as having lower impulsivity and positive responses to stimulation (adjusted mean [95% CI] “Surgency/Extraversion” in formula-fed vs. mixed-fed vs. breast-fed groups: 4.3 [4.2–4.5] vs. 4.0 [3.8–4.1] vs. 4.0 [3.9–4.1]; p-heterogeneity = 0.0006), lower ability to regulate their own emotions (“Orienting/Regulation”: 5.1 [5.0–5.2], vs. 4.9 [4.8–5.1] vs. 4.9 [4.8–5.0]; p = 0.01), and higher emotional instability (“Negative affectivity”: 2.8 [2.6–2.9] vs. 3.0 [2.8–3.1] vs. 3.0 [2.9–3.1]; p = 0.03). Conclusions Breast and mixed-fed infants were rated by their mothers as having more challenging temperaments in all three dimensions; particular subscales included greater distress, less smiling, laughing, and vocalisation, and lower soothability. Increased awareness of the behavioural dynamics of breastfeeding, a better expectation of normal infant temperament and support to cope with difficult infant temperament could potentially help to promote successful breastfeeding.