Antiserum to Hp is specific and identifies Hephaestin protein in C57BL/6J and sla mice. (A-B) A full-length mouse Hp cDNA containing a C-terminal FLAG tag was transiently transfected into CHO cells (CHO-Hp). After 48 hours, clarified cell lysates were collected and used for immunoprecipitations with the monoclonal anti-FLAG antibody. Immunoprecipitates were separated by SDS-PAGE and blotted onto nitrocellulose. The blots were probed with anti-FLAG (A) or anti-Hp (B). (C) Enterocyte extracts from C57BL/6J mice and sla mice were used for immunoprecipitations with the anti-Hp antibody. Immunoprecipitates were separated by SDS-PAGE, blotted, and probed with the anti-Hp antibody. An Mr 155 000 protein is present in the C57BL/6J, whereas an Mr 130 000 protein is present in the sla mice. Because less Hp protein is present in the sla mouse, we loaded 5 times more immunoprecipitate from the sla mouse to visualize the protein.
