| Objective: To study isolation and purification of rat spermatogonia and effects of bisphenol A on activities and morphology and structure of spermatogonia in vitro. Methods: Two digestions of three enzymes were used to prepare germ cell suspension of Wister rat at 10 days; Percoll discontinue density gradient centrifugation and differentiation of the velocity of cells sticking to the wall in vitro were used to isolate and purify spermatogonia respectively; Morphological and structural features of spermatogonia in rat at 10d of age were observed by light and transmission electron microscope; Spermatogonia was identified by c-kit immunochemistry. To study effects of the different dosages of bisphenol A ( 1× 104mol/L, 1×105mol/L, 1 × 10-6mol/L, 1 × 107mol/L) on spermatogonial activities in vitro; To observe morphological and structural transformation of the different dosages of bisphenol A on spermotogonia by stained with Giemsa. Results : Spermatogonia were mainly distributed in percoll gradient between 28%~36%, Their morphological and structural features were consistent with that in testis sections of 10 days old rat. Purification of spermatogonia was 55.68% after purified. Spermatogonia were c-kit—positive or strong positive cells, but c-kit expression of testic body cells was weak positive or negative. Bisphenol A could reduce the activities of spermatogonia in vitro, increase the number of nuclear solidification and shrinkage of spermatogonia in vitro, transform morphology and structure of spermatogonia in vitro. Conclution: Spermatogonia isolated and purified from rat at 10 days by two digestions of three enzymes, percoll discontinue density gradientcentrifugation and differentiation of the velocity of cells sticking to the wall in vitro could satisfy the needs of test in vitro . spermatogonia expressed specific c-kit receptor, bisphenol A could cause immediate toxicity on spermatogonia.
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