Cryoprotectant‐free vitrification of donkey sperm using 0.25 ml straws has been recently developed, but the obtained results have not been directly compared to conventional slow freezing yet. The aim of this study was to compare sperm quality parameters after cryopreservation using both methods. Semen samples were collected from three Andalusian Donkeys. Semen was centrifuged and pellets resuspended with an extender with glycerol for conventional freezing or the same extender without glycerol, but with sucrose 0.1 mol/L for vitrification. Conventional freezing was performed in nitrogen vapours and thawed in a water bath (30s/37°C). Vitrification was performed in covered 0.25 ml straws plunged directly into liquid nitrogen and warmed in 3 ml of a milk‐based extender at 43°C. Total (TM, %) and progressive motility (PM, %) were evaluated by computer‐assisted sperm analysis, and plasma membrane (PMI, %) and acrosome (AIS, %) integrities by epifluorescence microscopy. No differences (p > .05) were found between slow freezing and vitrification methods for any of the parameters assessed: TM (58.2 ± 16.1% vs. 52.7 ± 15.6%), PM (44.7 ± 18.2% vs. 44.3 ± 15.0%), PMI (55.4 ± 9.0% vs. 49.2 ± 11.2%) and AIS (38.4 ± 19.6% vs. 45.0 ± 11.0%), respectively. In conclusion, donkey sperm vitrification in straws presented similar sperm quality after thawing in comparison to conventional freezing. Therefore, it could be considered as an alternative to slow freezing regarding the sperm parameters assessed.

中文翻译：

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使用吸管玻璃化驴精子作为传统慢速冷冻的替代方法。

最近开发了使用 0.25 ml 吸管对驴精子进行无冷冻保护剂的玻璃化冷冻，但尚未将所获得的结果与传统的慢速冷冻直接进行比较。本研究的目的是比较两种方法冷冻保存后的精子质量参数。从三头安达卢西亚驴身上采集精液样本。将精液离心并用含有甘油的增量剂重新悬浮颗粒以进行常规冷冻，或者用不含甘油但含有 0.1 mol/L 蔗糖的相同增量剂进行玻璃化冷冻。常规冷冻在氮气蒸汽中进行，并在水浴 (30s/37°C) 中解冻。玻璃化是在有盖的 0.25 毫升吸管中进行的，吸管直接浸入液氮中，并在 43°C 的 3 毫升牛奶补充剂中加热。总 (TM, %) 和进行性运动 (PM, %）通过计算机辅助精子分析评估，质膜（PMI，%）和顶体（AIS，%）完整性通过落射荧光显微镜进行评估。无差异（p  > .05) 发现在慢速冷冻和玻璃化方法之间评估的任何参数：TM (58.2 ± 16.1% vs. 52.7 ± 15.6%), PM (44.7 ± 18.2% vs. 44.3 ± 15.0%), PMI (分别为 55.4 ± 9.0% 对 49.2 ± 11.2%）和 AIS（38.4 ± 19.6% 对 45.0 ± 11.0%）。总之，与传统冷冻相比，吸管中的驴精子玻璃化在解冻后呈现出相似的精子质量。因此，对于评估的精子参数，它可以被视为慢速冷冻的替代方案。