中文摘要：

【目的】主要探讨水牛腔前卵泡体外培养的影响因素。【方法】采用梳刮法从沼泽型水牛的卵巢中回收得到腔前卵泡,而后用McCoy＇s5a培养液在96孔培养板中体外培养5～10d,在显微镜下观测腔前卵泡的形态与增长情况,并用台盼蓝染色鉴定腔前卵泡的存活情况。【结果】在培养液中添加100μg·L^-1的FSH能显著提高腔前卵泡培养5和10d后的增长幅度,当同时添加50mg·L^-1的维生素C时增长幅度进一步加强;添加50mg·L^-1的维生素C能提高腔前卵泡培养10d后的形态正常率,但对卵泡增长及存活没有影响;添加50μg·L^-1的EGF能提高腔前卵泡培养5d的增长幅度,但对培养10d后腔前卵泡的作用不显著。随着卵泡的直径增加,体外存活率升高,增长的幅度上升,异常率下降。60～100μm的腔前卵泡用三维培养法（琼脂糖包埋）体外培养10d的直径增长幅度显著高于二维培养法（琼脂糖铺底）和普通培养法,且形态异常显著低于二维培养法和普通培养法;100～140μm卵泡用二维培养法培养的增长幅度最大,培养成活率最高。【结论】FSH能促进水牛卵泡的体外增长,维生素C能维持卵泡的正常形态结构,且两者的协同作用显著;水牛卵泡体外发育能力随其体积的增大而增强;三维培养法适宜于培养60～100μm的卵泡,而二维培养法有利于100～140μm卵泡的体外发育。

英文摘要：

[Objective] Factors affecting the in vitro development of buffalo preantral follicles were examined in this study. [Method] Buffalo preantral follicles isolated by Comb-scrape method were cultured in the McCoy＇s5a medium in the 96 well cell culture plate for 5 d to 10 d. Then, the growth and morphology of preantral follicles were evaluated under microscope and the viability of preantral follicles was identified by Trypan Blue staining. [Result] The growth of preantral follicles was stimulated by addition of 100 μg.L^-1 FSH to the medium at either 5 or 10 d of in vitro culture （IVC）, and this stimulating effect was enhanced in the presence of 50 mg.L^-1 ascorbic acid. Addition of 50 mg.L^-1 ascorbic acid to the medium could increase the proportion of preantral follicles with normal morphology shape after 10 d of IVC, but did not affect their growth. Supplementation of medium with 50 μg.L^-1 EGF stimulated the growth of preantral follicles at 5 d of IVC, but this beneficial effect was not shown at 10 d of IVC. As the follicular diameter increased, the survival rate and growth of follicles were increased, and the morphological abnormity of follicles was decreased. The growing extent of preantral follicles at diameter of 60-100 μm cultured in three dimension （agarose embed） culture system was significantly higher than that in conventional or two dimension （agarose coating） culture system, and the abnormity rate of follicles was lower after 10 d of IVC in comparison with other culture systems. However, the growing extent and survival rate of preantral follicle at diameter of 100-140 μm was the highest after 10 d of IVC in the two dimension culture system. [Conclusion] FSH can promote the growth of buffalo preantral follicles, and ascorbic acid is beneficial to the maintenance of follicular morphology. The growing ability of buffalo preantral follicles is increasing with their diameter at IVC. Three dimensions culture system is suitable for culturing the follicles at diameter of 60-100μm, while the