en تخصصي Special پژوهشي Original article <div><strong>Background and Aims:</strong> <em>Citrus</em> <em>tristeza</em> virus (CTV) is the most economically important pathogen of <em>citrus</em> throughout the world. To avoid the ominous effect of disease in <em>citrus</em> growing area, producing of virus free plants and elimination of infected plants are imperative. For this aim obtaining simple and sensitive diagnosis tools is crucial. The main objective of present study is applying of recombinant protein technology for production of specific antibody and developing of serological <em>assays</em> for efficient detection of CTV within infected plants.<br> <strong>Materials and Methods:</strong> The major coat protein (p25) of CTV was selected as a target for preparation of polyclonal antibody. The gene encoding p25 was recombinantly expressed in bacterial host and the protein was purified through affinity chromatography approach. The purified recombinant coat protein was used for immunization of rabbit. Specificity of the prepared serum against CP was confirmed through serological assay. The immunoglobulin molecules were purified from serum through staphylococcus protein A followed by conjugation to alkaline phosphatase (AP) and horse radish peroxidase (HRP) enzymes. The prepared antibodies and conjugates were used for detection of infected plants by double antibody sandwich- enzyme linked immunosorbent assay (DAS-ELISA) and dot-blot immune binding assay (DIBA).<br> <strong>Results:</strong> The p25 protein was expressed in bacterial host. The SDS-PAGE results confirmed high purity and integrity of CTV major coat protein with the expected size of about 29 kDa. The indirect ELISA results revealed that the antibody titer was around 1:65000. The IgG molecules purified through protein A column and SDS-PAGE results confirmed purity of the prepared antibody. The concentration of IgG was quantified by comparison to standard protein, BSA, which was estimated around 1 mg/ml.&nbsp; The results obtained from DAS-ELISA and DIBA assays proved that prepared antibodies could be effectively applied for detection of CTV infected plants. &nbsp;<br> <strong>Conclusions: </strong>The prepared antibody and conjugates were powerful tools for detection of infected plants. To the best of our knowledge this is the first work for applying of peroxidase enzyme in developing of ELISA assay against CTV.</div> Citrus tristeza virus, Polyclonal antibody, Recombinant protein, Serological assay. 8 15 http://journal.isv.org.ir/browse.php?a_code=A-10-193-1&slc_lang=en&sid=1 F Samiei fa_samiee@yahoo.com 10031947532846006747 10031947532846006747 No Department of Plant Pathology, Science and Research Branch, Islamic Azad University, Tehran, Iran MR Safarnejad mrsafarnejad@yahoo.com 10031947532846006748 10031947532846006748 Yes Department of Plant Viruses, Iranian Research Institute of Plant Protection, Agricultural Research Education and Extension Organization of Iran (AREEO), Tehran, Iran Gh Hosseini Salekdeh hsalekdeh@abrii.ac.ir 10031947532846006749 10031947532846006749 No Department of Genomics, Agricultural Biotechnology Research Institute of Iran, Karaj, Iran M Shams- Bakhsh shamsbakhsh@gmail.com 10031947532846006750 10031947532846006750 No Department of Plant Pathology, Faculty of agriculture, Tarbiat Modares University, Tehran, Iran. H Safarpour safarpour701@yahoo.com 10031947532846006751 10031947532846006751 No Cellular and Molecular Sciences Research Center, Birjand University of Medical Sciences, Birjand, Iran