Specificity of the localization of transforming growth factor-alpha immunoreactivity in colon mucosa

Transforming growth factor-alpha (TGF-alpha) plays an important role in gas trointestinal pathophysiology. However, the exact location of its expressio n in the intestine is still controversial. This study systematically compar ed the localization of TGF-alpha immunoreactivity in frozen or fixed human colon using three different antibodies and examined specificity of antibodi es by using tissues from TGF-alpha knockout mice and by Western blotting. C onsistent with the mRNA distribution revealed by in situ hybridization, a s imilar staining pattern was obtained in frozen sections by ail three antibo dies, localizing on the surface and along the crypt epithelium. In paraffin sections, although the polyclonal antibodies (raised against recombinant h uman or rat TGF-alpha) gave minimal staining, the monoclonal antibody (agai nst C-terminal peptide of human TGF-alpha) still gave intense staining on t he surface and upper crypt epithelium. By using specimens from TGF-alpha kn ockout mice in immunostaining and Western blotting, the polyclonal antibodi es were shown to be specific. In contrast, specificity of the monoclonal an tibody was in doubt in rodent tissues because it gave similar detection bet ween wild-type and knockout mice in both analyses, indicating its crossreac tion to non-TGF-alpha molecules. In conclusion, frozen sections and antibod ies raised from recombinant TGF-alpha should be used for TGF-alpha immunohi stochemistry in the colon.