TNF-⍺ is a principal pro-inflammatory cytokine which contributes to the activation of innate immunity and the transition to antigen-specific adaptive immunity in tuberculosis. Using recombinant guinea pig (rgp) TNF-⍺, the effect of TNF-⍺ on lymphocyte activation was examined in unvaccinated and BCG-vaccinated guinea pigs. Splenocytes were stimulated with PPD or ConA, in the presence or absence of rgp TNF-⍺ for 96h. Lymphocyte proliferation was measured using [³H]thymidine uptake, and IL-12 p40 and IFN-γ mRNA were analyzed using real-time PCR. rgpTNF-⍺ alone was able to stimulate a significant degree of proliferation in splenocytes. The addition of rgpTNF-⍺ to PPD-stimulated cells enhanced the proliferation of splenocytes from BCG-vaccinated guinea pigs. Furthermore, enhancement of proliferation by rgpTNF-⍺ was found to be correlated with upregulation of the levels of Type 1 cytokine mRNA (IL-12p40 and IFN-γ) in splenocyte cultures. This suggests that TNF-⍺ plays an important role in the regulation of Type 1 T cell-mediated immune responses in the guinea pig.