Rapid Assay of Cellular Immunity in Q Fever

Cellular of T lymphocyte function is important in defense against infectious disease. T lymphocytes also participate in tumor and transplant rejection, auto immune disease, and hypersensitivity. A central facet of T lymphocyte function is activation toward proliferation following exposure to specific antigen in the conted of the major histocompatibility complex on antigen presenting cells. We proposed to develop a rapid assay based om the large and early increase in ATP in quiescent cells that are activated using Q fever as a model system. Phase I or phase II antigen from C. brunetii, the causative agent for Q fever, will be added to whole blood samples diluted in buffer. After incubation, T lymphocytes or appropriate subsets will be separated and washed by immunomagnetic beads coated with either antibody to a marker found on all T lymphocytes or to antiboldy to CD4 or CD8. The lymphocytes will be disrupted and the ATP measured by addition of luciferin/luciferase. Antigen specific response will be measured relative to a buffer control and to a T cell mitogen. The proposed method offers a rapid, simple, and sensitive alternative to standard methods of monitoring T cell function. The potendtial sensitivity of the test is based on the large increase - upto 1000 fold - of ATP in stimulated cells and the high quantum efficiency of the luciferin-luciferase reaction. The method also offers the potential to measure activation of several subsets of T lymphocytes. After a prototype assay is developed, the test will be validated against standard tests and by study of patients with Q fever.