鸭胚睾丸支持细胞的分离培养与鉴定

为探究鸭胚睾丸支持细胞的分离培养方法，选取２４日胚龄的麻鸭鸭胚４０个，取出睾丸，采用胶原酶Ⅳ与胰蛋白酶二步消化分离睾丸细胞，差异贴壁与低渗处理的方法进行纯化，于３７℃，体积分数５％二氧化碳条件下培养，最终得到纯度较高的支持细胞．经染色鉴定，发现罗丹明１２３染色显示支持细胞中富含线粒体；ＡＫＰ检测发现８０％的细胞呈ＡＫＰ阴性；吖啶橙染色显示支持细胞细胞质中富含ＲＮＡ；油红Ｏ染色显示睾丸支持细胞细胞质中含有大量脂肪滴，细胞核内可见双极小体．本试验结果说明采用胶原酶Ⅳ与胰蛋白酶结合消化，经差异贴壁与低渗出处理纯化，可得到较高纯度的支持细胞；结合罗丹明１２３染色法、ＡＫＰ染色法、吖啶橙染色法与油红Ｏ染色法可有效鉴定支持细胞．

Isolation,culture and identification of testis sertoli cells from duck embryo

To explore the isolation and culture method of duck testis sertoli cells, a total of 40 duck embryos of 24 days old were selected in this experiment. Testis was taken out, and the testis cells were separated by two-step digestion with collagenase IV and trypsin, and then purified by differential adherence and hypotonic treatment. The cells were cultured at 37 ℃, 5% carbon dioxide, and finally high purity sertoli cells were obtained. After staining and identification, rhodamine 123 staining showed that the cells were rich in mitochondria. AKP test showed that 80% of the cells were AKP negative. Acridine orange staining showed that the cytoplasm of cells was rich in RNA. Oil red O staining showed that the cytoplasm of cells contained a large number of fat droplets and bipolar bodies were visible in the nucleus.The results of this experiment showed that high purity sertoli cells could be obtained by combining collagenase IV with trypsin for digestion and purification through differential adherence and hypotonic treatment. Combining rhodamine 123 staining, AKP staining, acridine orange staining and oil red O staining could effectively identify sertoli cells.