Salinity and drought are the most important environmental factors causing osmotic stress and limit intensely plant growth. Production of osmolyts such as proline is a current response in many plants under osmotic tension. The key enzyme of proline biosynthesis pathway is P5CS (D1-pyrroline- 5-carboxylate synthetase). In this study, embryos of the olive cv. Zard were cultured on solid 1/2 MS media, after 4 weeks, half of the plantlets were transferred to the solid 1/2 MS containing 200 mM NaCl and they were sub- cultured about 2 weeks, then total protein was extracted from fresh leaves of both plantlets under stress condition and non stress condition and measured using of brad ford to subject to SDS-PAGE gel electrophoresis. To probe the effect of P5CS overexpression on salinity stress tolerance, poly clonal antibody developed in rabbit against P5CS protein was used. The result of immunological methods for specific detection of P5CS gene, strongly indicated P5CS up-regulation in stressed plantlets versus non stressed plantlets and these results implicate the key role of proline in salinity stress toleration.