Objectives were to study the effect of storage for 3 wk at ‐20, 4, and 24°C on protein (TP), free amino ends (FAE), fat (F), free fatty acids (FFA), bacterial growth (B), secretory immunoglobulin A (SIgA) and lysozyme (L) activity in freshly expressed human breast milk (HM) and to examine bacterial growth in pasteurized (P) and unpasteurized (UP) HM in a bottle contaminated by an infant and stored at 24°C for up to 24 hr and at 4°C up to 6 days. HM samples were collected from 12 women and stored up to three weeks. There was a decrease in TP in HM stored at 24°C at 1 wk (p < 0.01), an increase in FAE at 4°C at 1 wk (p < 0.05) and 24°C at 1 wk (p<0.001), an increase in FFA at ‐20°C at 2 wk (p < 0.05), 4°C at 48 hr (p < 0.0001), and 24°C at 48 hr (p < 0.0001), and an increase in B at 4°C at 2 wk (p < 0.05) and 24°C at 48 hr (p < 0.01). HM samples were collected from 5 women whose infants partially consumed bottles of P and UP HM. There was an increase in B in P HM stored at 24°C at 24 hr (p < 0.0001) and in UP HM stored at 24°C at 24 hr (p < 0. 001). Infant suckling contributed less B than pumping to HM. The data indicate that HM may be safely stored longer than current recommendations.