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Expression of the high-affinity interleukin 2 receptor alpha chain (CD25) was used to monitor antigen-specific activation of T lymphocyte subsets (CD4+, CD8+, and gammadelta T cells) from cattle immunized with modified live bovine herpesvirus-1 (BHV-1). Calves seronegative for BHV-1 were either vaccinated with one dose of modified live vaccine containing BHV-1 or not vaccinated to serve as negative controls. Two animals vaccinated 7 and 5 weeks before the start of the experiment with two doses of modified live vaccine containing BHV-1 served as positive controls. Blood samples were taken from the nonvaccinate group, the positive control group, and the vaccinate group at 0, 21, 35, 60, and 90 days postinoculation (PI). Isolated peripheral blood mononuclear cells from immunized and control animals were incubated for 5 days with and without live BHV-1 ISU99. Compared to the nonvaccinates, a significant (p < 0.05) increase in expression of CD25 by CD4+, CD8+, and gammadelta T lymphocytes from the vaccinate group was detected following in vitro exposure to live BHV-1 after vaccination. This is apparently the first report using live BHV-1 to stimulate lymphocytes in vitro and showing CD8+ T cell activation. Peripheral blood from the positive control animals was depleted of CD4+, CD8+, or gammadelta T lymphocytes prior to incubation with BHV-1 to assess bystander activation in the CD25 expression assay. When incubated with live BHV-1, depletion of CD4+ T cells depressed the expression of CD25 by CD8+ T cells, but not gammadelta T cells. Depleting CD8+ or gammadelta T cells prior to in vitro culture with BHV-1 did not affect the expression of CD25 by the remaining T lymphocyte subsets. Vaccinates were protected from challenge with virulent BHV-1 at 110 days postvaccination compared to nonvaccinates. Expression of CD25 appears to be a useful marker for evaluating induction of antigen-specific T lymphocyte subset responses following vaccination.

Bovine peripheral blood mononuclear cells from cattle immunized with a modified live virus bovine herpesvirus 1 (BHV1) vaccine and from BHV1-negative cattle were incubated in vitro with inactivated BHV1 for 6 days. Activation of T-lymphocyte subsets was measured by two-color flow cytometric analysis of T-cell phenotype and surface expression of CD25, the alpha-subunit of the high-affinity interleukin-2 receptor. Vaccinated animals, but not unvaccinated animals, had CD3+, CD4+, and gamma-delta T cells that significantly (p < 0.05) increased expression of CD25 when incubated with BHV1. CD8+ T cells from vaccinated animals did not consistently increase CD25 expression when incubated with inactivated BHV1.

CHAPTER 1. GENERAL INTRODUCTION 1 Dissertation organization 1 Research summary 2 CHAPTER 2. LITERATURE REVIEW: IN VITRO DETECTION OF ANTIGEN SPECmC MEMORY T LYMPHOCYTES 4 Introduction: the importance of memory lymphocytes 4 What are memory lymphocytes? 5 Applications for detection of antigen specific memory lymphocytes 7 T lymphocyte subsets 10 Methods of detection in vitro of T cell activation 12 Summary 21 References 22 CHAPTER 3.