Background: Most preterm infants receive antibiotics to prevent serious infections shortly after birth. However, prolonged antibiotic treatment predisposes to gut dysbiosis and late-onset sepsis. Using preterm pigs as model, we hypothesized that neonatal prophylactic antibiotics impair systemic immune development beyond the days of antibiotic treatment. Methods: Preterm pigs (90% gestation) were fed formula for 9 days, treated with sterile water (CON) or enteral antibiotics from day 1 to 4. On days 5 and 9, blood was collected for haematology, in vitro LPS stimulation, and plasma proteomics. Results: Antibiotic treatment altered the abundance of 21 and 47 plasma proteins on days 5 and 9, representing 6.6% and 14.8% of the total annotated proteins, respectively. Most antibiotics-induced proteome changes related to complement cascade, neutrophil degranulation, and acute phase responses. Neutrophil and lymphocyte counts were higher in antibiotics-treated pigs on day 5 but did not change from days 5–9, in contrast to increasing cell counts in CON. The antibiotics treatment suppressed TNF-alpha and IL-10 responses to in vitro LPS challenge on day 5, 7 and 9. Conclusion: Few days of antibiotics treatment following preterm birth alter the plasma proteome and inhibit systemic immune development, even beyond the days of treatment. Impact: 1.Neonatal prophylactic antibiotics alter the plasma proteome and suppress systemic immune development in preterm pigs2.The effects of prophylactic antibiotics last beyond the days of treatment.3.Neonatal antibiotics treatment for compromised human newborns may predispose to longer-term risks of impaired immunity and infections.