This experiment was conducted to investigate the effect of iron source on Fe absorption and the gene expression of divalent metal transporter 1 (DMT1) and ferroportin 1 (FPN1) in the ligated duodenal loops of broilers. The in situ ligated duodenal loops from Fe-deficient broiler chicks (28-d-old) were perfused with Fe solutions containing 0 to 14.33 mmol Fe/L from 1 of the following: Fe sulfate (FeSO∙7HO), Fe methionine with weak chelation strength (Fe-Met W; chelation strength is expressed as quotient of formation [Q] value, Q = 1.37), Fe proteinate with moderate chelation strength (Fe-Prot M; Q = 43.6), and Fe proteinate with extremely strong chelation strength (Fe-Prot ES; Q = 8,590) for up to 30 min. The gene expression of DMT1 and FPN1 in the duodenal loops from the control group and the groups treated with 3.58 mmol Fe/L from 1 of 4 Fe sources was analyzed. The absorption kinetics of Fe from different Fe sources in the duodenum followed a saturated carrier-dependent transport process. The maximum transport rate (J) values in the duodenum were greater ( < 0.03) for Fe-Prot ES and Fe-Prot M than for Fe-Met W and FeSO∙7HO. The Fe perfusion inhibited ( < 0.05) the mRNA expression of but enhanced ( < 0.0008) the mRNA expression of in the duodenum and had no effect ( > 0.14) on the protein expression levels of the 2 transporters. These results indicated that organic Fe sources with greater Q values showed higher Fe absorption; however, all Fe sources followed the same saturated carrier-dependent transport process in the duodenum, and DMT1 and FPN1 might participate in Fe absorption in the duodenum of broilers regardless of Fe source.