ABSTRACT
<p><b>OBJECTIVE</b>To express and purify of the BC097361 recombinant protein, and to prepare the BC097361 specific rabbit polyclonal antibody.</p><p><b>METHODS</b>BC097361 cDNA was ligated into the prokaryotic expressive vector pET-32a (+), and the resulting plasmid was transformed into E.coli BL21 (DE3). The protein expression was induced with IPTG and the protein was analyzed with SDS-PAGE and western blotting. The expressed product was purified using Ni+ affinity column chromatography.Then the purified pET-32a (+) -BC097361 fusion protein was used to immunize New Zealand rabbits to gain polyclonal antibody. The specificity and potency of polyclonal antibody were evaluated by Western blot and ELISA.</p><p><b>RESULTS</b>The BC097361 fusion protein was highly expressed.The protein was mainly in the inclusion body. ELISA indicated the titer of polyclonal antibody more than 1:320000. The high specificity was confirmed with Western blot.</p><p><b>CONCLUSIONS</b>The recombinant BC097361 fusion protein and the BC097361 specific polyclonal antibody will be valuable tools for the investigation on the biological function of BC097361.</p>
Subject(s)
Animals , Male , Rabbits , Angiotensin II , Genetics , Antibodies , Allergy and Immunology , Metabolism , Antibody Specificity , Blotting, Western , Cloning, Molecular , Escherichia coli , Genetics , Metabolism , Gene Expression , Genetic Vectors , Genetics , Liver Cirrhosis , Genetics , Plasmids , Genetics , Recombinant Fusion Proteins , Allergy and Immunology , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
<p><b>OBJECTIVE</b>To detect TB specific T cell responses by using the recombinant ESAT-6 protein as stimulus in Chinese HIV infected patients.</p><p><b>METHODS</b>ELISPOT-IFN-gamma assay by using the recombinant ESAT-6 protein as stimulus to detect specific T cell responses in HIV+ patients with or without clinical manifestation of TB diseases.</p><p><b>RESULTS</b>Recombinant ESAT-6 protein specific T cell responses show significant high frequencies in both of TB patients with or without HIV infection than that in the healthy control and HIV+ group without clinical TB diseases.</p><p><b>CONCLUSION</b>The ELISPOT-IFN-gamma assay by using recombinant ESAT-6 protein as stimulus could be used in diagnoses of TB infection in Chinese HIV infected patients.</p>