Chemokine receptor 7 (CCR7)-expression and IFNγ production define vaccine-specific canine T-cell subsets

Canines suffer from and serve as strong translational animals models for many immunological disorders and infectious diseases. Routine vaccination has been a mainstay of protecting dogs through the stimulation of robust antibody responses and expansion of memory T-cell populations. Commercially available reagents and described techniques are limited for identifying and characterizing canine T-cell subsets and evaluating T-cell-specific effector function. To define reagents for delineating naïve versus activated T-cells and identify antigen-specific T-cells, we tested anti-human and anti-bovine T-cell specific cell surface marker reagents for cross-reactivity with canine peripheral blood mononuclear cells (PBMCs. Both CD4(+) and CD8(+) T-cells from healthy canine donors showed reactivity to CCL19-Ig, a CCR7 ligand, and coexpression with CD62L. An in vitro stimulation with concanavalin A validated downregulation of CCR7 and CD62L expression on stimulated healthy control PBMCs, consistent with an activated T-cell phenotype. Anti-IFNγ antibodies identified antigen-specific IFNγ-producing CD4(+) and CD8(+) T-cells upon in vitro vaccine antigen PBMC stimulation. PBMC isolation within 24h of sample collection allowed for efficienT-cell recovery and accurate T-cell effector function characterization. These data provide a reagent and techniques platform via flow cytometry for identifying canine T-cell subsets and characterizing circulating antigen-specific canine T-cells for potential use in diagnostic and field settings.