SummaryThe present study evaluated the effect of knockout serum replacement (KSR), fetal bovine serum (FBS) and bovine serum albumin (BSA) on the viability and growth of bovine secondary follicles culturedin vitrofor 12 days. To this end, secondary follicles were isolated (185–202 μm) and culturedin vitroin TCM-199+medium supplemented with KSR (5% and 10%), FBS (5% and 10%) or BSA (3 mg/ml) at 38.5°C with 5% CO2in air. Follicular diameters were evaluated on days 0, 4, 8 and 12. After 12 days of culture, follicular survival analysis was performing by using calcein-AM and ethidium homodimer. Before and after culture, follicles were fixed in paraformaldehyde for histological evaluation. Follicular diameter at different days of culture were compared using the Kruskal–Wallis test, while the percentages of viable follicles were analyzed by chi-squared test (P< 0.05). Results showed that follicles cultured in the presence of KSR at both concentrations presented higher follicular survival rates than those cultured in control medium alone or supplemented with FBS or BSA. Conversely, the presence of KSR, BSA or FBS did not increase follicular diameter after 12 days of culture. Histology analysis showed that, among the tested treatments, follicles cultured in the presence of KSR had preserved rounded oocytes, juxtaposed granulosa cells and intact basal membrane. In conclusion, supplementation of culture medium with KSR increases the follicular survival of bovine secondary follicles culturedin vitro.

中文翻译：

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与体外培养过程中的其他蛋白质来源相比，补充具有敲除​​血清替代物的培养基可提高牛次级卵泡的存活率

摘要本研究评估了敲除血清替代物 (KSR)、胎牛血清 (FBS) 和牛血清白蛋白 (BSA) 对培养的牛次级卵泡的活力和生长的影响体外12天。为此，分离次级卵泡（185-202 μm）并培养体外在中医-199+培养基中添加了 KSR（5% 和 10%）、FBS（5% 和 10%）或 BSA（3 mg/ml），温度为 38.5°C，含 5% CO2在空气中。在第 0、4、8 和 12 天评估卵泡直径。培养 12 天后，使用 calcein-AM 和 ethidium homodimer 进行卵泡存活分析。培养前后，将卵泡固定在多聚甲醛中进行组织学评估。使用Kruskal-Wallis检验比较不同培养日的卵泡直径，而通过卡方检验分析存活卵泡的百分比。磷< 0.05)。结果表明，在两种浓度的 KSR 存在下培养的卵泡比在单独或补充 FBS 或 BSA 的对照培养基中培养的卵泡存活率更高。相反，KSR、BSA 或 FBS 的存在在培养 12 天后并未增加卵泡直径。组织学分析表明，在测试的处理中，在 KSR 存在下培养的卵泡保留了圆形卵母细胞、并列的颗粒细胞和完整的基底膜。总之，在培养基中添加 KSR 可增加培养的牛次级卵泡的卵泡存活率体外.