Breast-fed preterm infants often show a better outcome, partly ascribed to the benefit of insulin-like growth factors (IGFs) and their binding proteins (IGFBP). We compared IGF-I, IGF-II, IGFBP-2 and IGFBP-3 levels, measured by radioimmunoassays in milk samples from 30 mothers of preterm (<31 weeks) and from 19 mothers of term (>37 weeks) infants at days 7 and 21 postpartum. Proteolysis of IGFBP-2 within mother’s milk and digestion of ¹²⁵I-IGF-II and ¹²⁵I-IGFBP-2 by gastric juice from neonates were assessed by electrophoretic techniques. Mean concentrations did not differ between preterm and term milk: IGF-I (2.8 � 0.2 vs. 2.3 � 0.1 ng/ml), IGF-II (12.0 � 0.4 vs. 12.2 � 0.5 ng/ml), IGFBP-3 (100.0 � 5.1 vs. 80.0 � 5.8 ng/ml), but did so for IGFBP-2 (3,144 � 172 vs. 2,428 � 188 ng/ml, p < 0.02). Immunoblots revealed 42% (p < 0.05) more IGFBP-2 fragments of 14 and 25 kDa in preterm milk. Incubation with gastric juice caused cleavage of ¹²⁵I-IGFBP-2 and partial cleavage of ¹²⁵I-IGF-II. Mutual complexation protected IGF-II and IGFBP-2 from cleavage, suggesting that both are likely to arrive in the bowel in an intact form to exert promotive effects. The results provide further evidence that IGFBP-2 and IGF-II in breast milk are relevant factors for the early development of preterm infants.