Major subpopulations of gamma delta T cells within ruminant and pigs are defined by expression of WC1, a hybrid pattern recognition receptor/co-receptor to the T cell receptor (TCR). It is known that when WC1 is knocked down cells fail to respond. Showing that WC1 plays an active role in the stimulation of bovine gamma delta T cells. Here we explored the spatio-temporal dynamics of WC1 and TCR interaction using imaging flow cytometry and stochastic optical reconstruction microscopy. We found that in quiescent gamma delta T cells both WC1 and TCR existed in separate protein domains (protein islands) but after activation using Leptospira, our model system, that they concatenated. In cattle, WC1+ gamma delta T cells have been shown to use TCR gamma genes from only one of the six available cassettes (TRGC5). We postulated that this structure may be necessary to interact with WC1 for signal transduction. If correct, other species should have the same restriction of their T cell receptor (TCR) gene usage by their WC1+ cells. When evaluated by RT-PCR and PacBio sequencing we found that caprine WC1+ gamma delta T cells exhibited the same restriction as found in cattle while porcine WC1+ gamma delta T cells used all TCR gamma C genes, although they preferentially expressed TCR chains from their TRGC5 homologue cassette. Next, we addressed WC1 and TCR roles of determining antigen specificity. One model is that co-ligation of WC1 and the gamma delta TCR by antigen increases a low affinity gamma delta TCR-antigen interaction. In this paradigm, WC1 is the main determining element regarding pathogen recognition. If correct, we predict the TCR CDR3 sequences in the responding gamma delta T cells to be relatively unrestricted. The alternative model is that gamma delta T cells that respond to a pathogen have a TCR with higher affinity to antigen and thus would be restricted while TCR expressed on cell that do not respond would be polyclonal. To test these models, we performed next generation sequencing of sorted antigen-responding and non-responding WC1+ gamma delta T cells. As baseline data, populations of responding and nonresponding WC1+ gamma delta T cells were also evaluated for their overall transcriptome differences using RNA-Seq.