Forty-one normal horses were evaluated for reactivity to intradermally injected aqueous allergens to determine allergen threshold concentrations (TC), with potential relevance to equine intradermal testing (IDT). Horses were tested three times over 1 year to assess seasonal variation in reactivity, using three to five serial dilutions of 27 allergens each time. Injection sites were evaluated after 15 min, 1 h, 4 h and 24 h. The highest allergen concentration at which < 10% of horses demonstrated positive reactivity (subjective score of > or = 2, scale of 0 to 4) at 15 min was considered the TC. The TC was determined for nine pollens (2000 to > 6000 PNU mL(-1)), four moulds (4000 to > 6000 PNU mL(-1)), seven insects (ant, horse fly 125 PNU mL(-1); house fly, cockroach 250 PNU mL(-1); moth 60 PNU mL(-1); mosquito 1000 PNU mL(-1); Culicoides nebeculosis 1 : 5000 w v(-1)) and three of four storage mites (1 : 10,000 w v(-1)). The TC was not determined due to excessive reactivity at the lowest concentrations tested for dust mites (Dermatophagoides farinae [< 1 : 12,000 w v(-1)], D. pteronyssinus [< 1 : 30,000 w v(-1)]), and Acarus siro (< 1 : 10,000 w v(-1)). Minor variation in the TC for specific allergens occurred in different seasons. Progressive sensitization with repeat testing occurred for grain mill dust mix. Positive reactivity at 1 h and 4 h occurred in > 10% of horses for nine of 19 allergens (pollens, mosquito, storage mites) at their determined TC. Positive reactivity was rare at 24 h. This study in normal horses suggests that appropriate testing concentrations of allergens for equine IDT in atopic horses may be > or = 1000 PNU mL(-1) for pollens and moulds, 60 to 250 PNU mL(-1) for most insects and < 1 : 12,000 w v(-1) for dust mites; and that reactions at 1-4 h may be insignificant.