Four hundred and twenty sheep were examined for Oestrus ovis larval infestation by naked eye and serological test (Direct ELISA) during six months (from June to November, 2005). Excretory - secretory product (ESP) from first (L1) and second (L2) instar of O.ovis larvae were used as a coating antigen and a vaccine. The naked eye examination showed a positive prevalence of 19.5 % while the ELISA test showed 24.3 %. 251 Assuit Vet. Med. J. Vol. 52 No. 109 April 2006 The obtained data proved that the sensitivity of ELISA test was 97.6 %. Twenty female lambs of three months old were allocated into two groups. The first one received two IM injections of ESP in the neck, 4 weeks apart, initially in Freunds complete adjuvant and then in Freunds incomplete adjuvant. In control group, the animals received two injections of PBS with Freunds complete and incomplete adjuvants. Challenge test was carried out twice, 15 (15 larvae per animal) and 28 days (25 larvae per animal) after the second immunization. Sera samples were collected for ten weeks. On the 5th week post the second challenge test, all animals were slaughtered, all larvae were collected, identified and counted. ELISA data showed that O.ovis antibody began to increase one week after the first immunization and reached a peak on the 4" week post the second dose. Two animals among control group showed a moderate level of antibody, one on the 5th week and other on the 8th week. The results of challenge test showed that the establishment rate of O.ovis larvae were 33.5 % (134 out of 400) and 15 % (60 out of 400) in the control and vaccinated group respectively. It was concluded that diagnosis of O.ovis by ELISA using L1 and L2 ESP as a coating antigen is considered effective and the results obtained with O.ovis ESP L1 and L2 immunization are encouraging.