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This study examines differences in gender among entrepreneurs engaged in E-business activities. The empirical research was conducted in Thessaly, using a structured questionnaire. The 84 male and female who responded to our survey, were owners of micro enterprises. Results reveal differences in gender regarding E-business adoption. Males consider security and unnecessary adoption are more important barriers, while females believe E-business is not included in firm's strategic planning. Results also highlight the importance of awareness, consulting and development of specific skills by entrepreneurs, as barriers of E-business adoption.