The present experiment was initiated in order to study the immunological aspects of the etiology of gingival hyperplasia. To confirm that Phenytoin (Ph) it self can provoke an immune response as partial immunogen in the animal body, the least modified compound of Ph (p-aminophenytoin) was used for bridging with carrier protein (BSA). Rabbits were immunized initially with BSA-Ph in Freund's complete adjuvant and booster injections were carried out with BSA-Ph in Freund's incomplete adjuvant. The radio-immunoassay was carried out by competing unlabelled Ph with a known amount of 3H Ph, and B/T % was measured. The assay can detect as little as 2 μg of Ph. The specificity reaction of antisera with Ph metabolite or the other anticonvulsants revealed that antisera reacted with HPPH(major metabolite of Ph) very slightly but not at all with other anticonvulsants. Parallel analysis performed with HPLC with the same sample revealed excellent correlation between the two methods. The method here described is very simple and provides high accuracy. The success of anti-Ph antisera formation with antigen which provides unmodified Ph as the hapten seems to ensure that BSA-Ph conjugate can be used as a suitable immunogen in provocation of delayed hypersensitivity of Ph.