Baseline information on the functional responses expected for assays used to assess immunocompetence in the Northern bobwhite (Colinus virginianus) are largely unavailable. Our primary objective was to develop an in vitro lymphoproliferative response assay for assessing cell-mediated immunocompetence in the Northern bobwhite. Culture conditions were optimised for domestic Northern bobwhites and field tested on splenocytes from wild-caught quail. Results indicated that increasing cell concentration and media volume in culture, as well as decreasing concentrations of serum in media, improved splenocyte responses to Con A stimulation. Optimum culture conditions were attained with 1 million cells per well cultured in 200 μl of AIM-V serum-free media for 72 h. Five micrograms concanavalin A (Con A) or 2.5 μg Salmonella typhimurium mitogen (STM) per well provided maximum stimulation as measured by 3H-thymidine incorporation. Stimulation indices of splenocyte cultures of wild-caught Northern bobwhites to 5 μg Con A were approximately four-fold greater than levels observed for domestic quail (P=0.0055). Alternatively, stimulation indices of splenocyte cultures obtained from wild-caught and domestic Northern bobwhites to 2.5 μg STM per well were not different (P=0.3938).