A human ileal lipid-binding protein (ILBP3) cDNA was isolated and shown to encode a 128 amino acid protein with homology to the fatty acid-binding protein gene family. The human ILBP amino acid sequence exhibited 78% identity to the rat ILBP sequence. Northern blot analysis revealed a single transcript of approx. 0.6 kb in small intestine, but no hybridization to liver, spleen, thymus, prostate, ovary, caecum, or colon. Southern blot analysis of genomic DNA from a panel of rodent-human somatic cell hybrids revealed that the human ILBP gene resides on chromosome 5 and is not linked to any other known fatty acid-binding protein family member.