Development and optimization a high sensitive and specific ELISA system for rapid detection of paratuberculosis in cattle
Journal Title: International Journal of Advanced Biotechnology and Research. - Year 2016, Vol 7, Issue 1
Abstract
ABSTRACT: John’s disease is a chronic gastroenteritis of cattle which caused by Mycobacterium avium subsp. Paratuberculosis and has a wide spread all over the world. This bacterium can be found in feces and milk and transmits the disease easily. Therefore, rapid detection of infection is highly important. There are different ways for detection of paratuberculosis that ELISA has the highest importance between conventional methods because of its convenience and high accuracy. The aim of this study is development and optimization an ELISA system for rapid detection of paratuberculosis in cattle. For this purpose, secreted antigens of Mycobacterium avium subsp. Paratuberculosis by trichloroacetic acid (TCA) method were precipitated and the best concentration of antigen and antibody were determined by checkerboard and the system’s cut-off was determined with RUC curve; The specificity and sensitivity of developed ELISA system were evaluated for 1000 serum samples. In this study, the best concentration of antigen and antibody were 1.1 µg/l and 0.01 respectively and based on RUC curve results the amount of cut-off was evaluated 0.2 (95% CI, 82.94 to 99.91). Finally, specificity and sensitivity of system were determined 99% and 86% respectively that this system was more sensitive in comparison with IDEXX ELISA kit. These results showed that secreted antigens of Mycobacterium avium subsp. Paratuberculosis can be the best choice for developing new ELISA systems because the most present kits are using cellular antigens. Key Words: ELISA-paratuberculosis - checker board- IS900 PCR- ROC
Authors and Affiliations
Keshavarz Rouhollah* , Mirjalili Ali, Mosavari Nader, Tadayon Keyvan, Ghaemmaghami Shamseddin, Fardid Farzaneh and Dashti pour s
Keywords
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