<p>Background: The aim of the present study was to evaluate the effectiveness of human ovarian vitrification protocol followed with <em>in vitro</em> culture at the morphological and molecular levels.<br /> Methods: Ovarian tissues were obtained from 10 normal transsexual women and cut into small pieces and were divided into non-vitrified and vitrified groups and some of the tissues fragments in both groups were randomly cultured for two weeks. The morphological study using hematoxylin and eosin and Masson&#39;s trichrome staining was done. The analysis of mean follicular density, 17-&beta; estradiol (E2) and anti mullerian hormone (AMH), and real-time RT-PCR was down for the evaluation of expression of genes related to folliculogenesis. Data were compared by paired-samples and independent-samples T test. Values of p&lt;0.05 were considered statistically significant.<br /> Results: The proportion of normal follicles did not show significant difference between vitrified and non-vitrified groups before and after culture but these rates and the mean follicle density significantly decreased in both cultured tissues (p&lt;0.05). The expression of genes was similar in vitrified and non-vitrified groups but in cultured tissues the expression of GDF9 and FSHR genes increased and the expression of FIGLA and KIT-L genes decreased (p&lt;0.05). An increase in E2 and AMH concentration was observed after 14 days of culture in both groups.<br /> Conclusion: In conclusion, the present study indicated that the follicular development and gene expression in vitrified ovarian tissue was not altered before and after <em>in vitro</em> culture, thus this method could be useful for fertility preservation; however, additional studies are needed to improve the culture condition.</p>