Although documented stability of allergens used for diagnosis is important, research in this area has been limited. Most studies on extract stability have been of limited duration and discrepancies have been reported between stability test results of in vivo and in vitro methods. In this study we determined the stability of allergenic extracts, comparing the intracutaneous test and enzymallergosorbent test inhibition method and determining the effect of temperature, dilution, and preservatives. Three formulations of timothy pollen, birch pollen, house dust mite (D. pteronyssinus) and cat dander extracts, as used for bronchoprovocation, skin prick testing and intracutaneous testing, were stored for 24 months at 6 degrees centigrade. The influence of temperature on various formulations was determined using the enzymallergosorbent test inhibition technique during storage for up to 36 months. Most formulations were found to be stable for 24 (intracutaneous test) or 36 (enzymallergosorbent test inhibition) months at 6 degrees centigrade. At 25 degrees centigrade, most formulations showed a decrease in relative potency, which remained above the limit of 0.3 times the in-house-reference for the bronchoprovocation formulation of timothy pollen, birch pollen, and house dust mite and for the skin prick test formulation of cat dander. Cat dander was remarkably stable at 6 and 25 degrees centigrade in glycerine and birch pollen was very susceptible to phenol. This destructive effect of phenol could be prevented by adding human serum albumin. The discrepancy between in vivo and in vitro tests reported by others was confirmed for house dust mite and timothy pollen.