Expression of the recombinant EhCFIm25.

<p>A) Expression of the 6x-His-labeled EhCFIm25 protein. Bacteria <i>E. coli</i> were transformed with pRSET<i>-EhCFIm25</i> plasmid and protein expression was induced by the addition of 1 mM IPTG for 3 h. Proteins extracts (30 µg) were separated through 10% SDS-PAGE and gels were stained with Coomassie blue. Lane 1, molecular weight; lane 2, non-induced bacterial extract; lane 3, IPTG-induced bacterial extract. B) Immunodetection of rEhCFIm25 polypeptide by Western blot assays using anti-6x His tag antibodies. Lane 1, non-induced bacterial extract (30 μg); lane 2, IPTG-induced bacterial extract (30 μg). C) Purification of the 6x-His-labeled EhCFIm25 protein through affinity chromatography using a Ni-NTA column. Lane 1, molecular weight; lane 2, IPTG-induced bacterial extract; lane 3, unbound proteins; lane 4, wash using 150 mM imidazole; lanes 5-8, elution with 250 mM imidazole. D) Immunodetection of rEhCFIm25 polypeptide by Western blot assays using specific rabbit antibodies anti-EhCFm25Im. Lane 1, non-induced bacterial extract; lane 2, IPTG-induced bacterial extract; lane 3, IPTG-induced bacterial extract and anti-6x His tag antibodies used as control. E) Immunodetection of purified rEhCFIm25 polypeptide by Western blot assays. Lane 1, anti-6x His tag antibodies; lane 2, specific rabbit antibodies anti-EhCFIm25; lane 3, pre-immune serum; lane 4, control without primary antibody. Arrowhead, EhCFIm25.