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Title: Application of long-term cultured interferon-gamma enzyme-linked immunospot assay for assessing effector and memory T cell responses in cattle

item MAGGIOLI, MAYARA - Iowa State University
item Fosse, James
item Palmer, Mitchell
item VORDERMEIER, H - Veterinary Laboratories Agency (VLA)
item WHELAN, ADAM - Veterinary Laboratories Agency (VLA)
item Nonnecke, Brian
item Waters, Wade

Submitted to: Journal of Visualized Experiments
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/23/2015
Publication Date: 7/11/2015
Citation: Maggioli, M.F., Fosse, J.M., Palmer, M.V., Vordermeier, H.M., Whelan, A.O., Nonnecke, B.J., Waters, W.R. 2015. Application of long-term cultured interferon-gamma enzyme-linked immunospot assay for assessing effector and memory T cell responses in cattle. Journal of Visualized Experiments. (101):e52833. doi: 10.3791/52833.

Interpretive Summary: Despite highly successful eradication efforts in several countries, tuberculosis of cattle remains a serious health concern worldwide. In addition, an outbreak of tuberculosis in white-tailed deer in Michigan and continued importation of tuberculous cattle from Mexico have seriously hindered eradication efforts within the United States. Without new strategies, eradication and control of bovine tuberculosis will be impossible. Thus, improved techniques for control, such as better diagnostic tests and vaccines, are needed for prevention of tuberculosis in cattle. In this article, a technique is described using written and audiovisual formats. The technique is for an assay to determine immune responses that correlate with protective responses to bovine tuberculosis. Thus, it is crucial for bovine tuberculosis control efforts as it enables prioritization of vaccine candidates for costly efficacy studies. Knowledge described in this study will assist in the control of tuberculosis in cattle.

Technical Abstract: Effector and memory T cells are generated through developmental programing of naïve cells following antigen recognition. If the infection is controlled, up to 95% of the T cells generated during the expansion phase are eliminated (i.e., contraction phase) and memory T cells remain, sometimes for a lifetime. In humans, two functionally distinct subsets of memory T cells have been described based on the expression of lymph node homing receptors. Central memory T cells express C-C chemokine receptor 7 and CD45RO and are mainly located in T-cell areas of secondary lymphoid organs. Effector memory T cells express CD45RO, lack C-C chemokine receptor 7 and display receptors associated with lymphocyte homing to peripheral or inflamed tissues. Effector T cells do not express either CCR7 or CD45RO but upon encounter with antigen produce effector cytokines, such as interferon-gamma. Interferon-gamma release assays are used for the diagnosis of bovine and human tuberculosis and detect primarily effector and Effector memory T cell responses. Central memory T cell responses by CD4+ T cells to vaccination, on the other hand, may be used to predict vaccine efficacy, as demonstrated with Simian Immunodeficiency Virus infection of non-human primates, tuberculosis in mice, and malaria in humans. Several studies with mice and humans as well as unpublished data on cattle, have demonstrated that interferon-gamma ELISPOT assays measure central memory T-cell responses. With this assay, peripheral blood mononuclear cells are cultured in decreasing concentration of antigen for 10 to 14 days (long-term culture), allowing effector responses to peak and wane; facilitating central memory T cells to differentiate and expand within the culture.