Antigen E (AgE), the principal allergen of short ragweed pollen, has previously been quantitated by histamine release from sensitized leukocytes or by immunodiffusion. To measure AgE activity a double antibody radioimmunoassay (RIA) was developed. With this assay as little as 7 ng. of AgE was measurable, and by logit transformation of inhibition curves AgE activity in various allergy extracts could be compared immunologically. AgE reactivity was detected in extracts of several Ambrosia and Franseria species, but only AgE from short ragweed produced inhibition curves parallel to AgE IV-C. With giant ragweed this cross-reactivity was confirmed with immunodiffusion analysis. Certain lots of extracts from botanically unrelated pollens contained small amounts of AgE with inhibition curves parallel to AgE from short ragweed. No dialyzable AgE activity was found in freshly prepared short ragweed extracts. Extraction of short ragweed pollen with 50. per cent glycerin yielded extracts with greater AgE content than extraction with Coca's solution. AgE activity from short ragweed survived freezing and thawing (10 times) and was stable for months at 4 °C. in concentrated aqueous solutions. The quantities of AgE coupled to solidphase polymers used in the radioallergosorbent test could also be estimated with RIA.