Background: Environmental exposure chambers (EECs) have been used extensively to study allergic rhinoconjunctivitis. Few studies have been published using EECs in conjunctivitis only, and none have used conjunctival allergen challenge as a selection criterion. The present study validated ALYATEC EEC in allergic conjunctivitis to birch allergens. Methods: Sixteen patients with a positive conjunctival allergen challenge (CAC) were ex-posed to 60 ng/m3 of Bet v 1 in an EEC on 2 consecutive days for a maximum of 4 hours. Re-producibility was tested among seven of the patients. A positive conjunctival response during the CAC and the EEC exposure was defined as a Total Ocular Symptom Score (TOSS) ≥ 5. Results: Fifty percent of patients had a positive conjunctival response during the first expo-sure and 75% during the second. The mean time to a positive conjunctival response was 81.2±33.9 minutes and 101.6±57 (P>0.05) during the first and second exposure, respectively. No difference in the TOSS occurred between the two exposures. The time necessary to ob-tain a positive response during the CAC was significantly shorter than with the EEC. The es-timated quantity of Bet v 1 inducing a positive response was 0.07±0.03 ng (exposure 1), 0.07±0.07 ng (exposure 2), 980±784 ng (CAC). The frequency of conjunctival responses and quantity of Bet v 1 was reproducible in all six EEC exposures. Conclusions: Birch allergen exposures inducing early conjunctival responses were different than those identified with direct installation during CAC. EEC appears to be closer to natural exposure than CAC.