A practical in vitro sperm-egg binding assay that detects subfertile males
A practical in vitro sperm-egg binding assay that detects subfertile males
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DOI：
10.1095/biolreprod58.3.686
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发表时间：
1998-03-01
影响因子：
3.6
通讯作者：
Hammerstedt, RH
中科院分区：
文献类型：
--
作者：
Barbato, GF;Cramer, PG;Hammerstedt, RH
开发了一系列精卵结合的体外试验来识别潜在的生育力低下的公鸡。初始试验使用置于显微镜载玻片上的完整鸡蛋卵黄周膜（PVM）片段或在平底微孔板内干燥的PVM热溶解提取物（MS-PVM），用DNA特异性染色和落射荧光显微镜检测结合的精子。本研究建立了一种自动化的精子结合率测定方法，采用预先染色的磺基琥珀酰亚胺基-7-氨基-4-甲基香豆素-3-乙酸酯-HS-PVM，用荧光酶标仪计数结合精子。对4个不同生育力鸡群进行了测定，结果表明：1）精子结合率与生育力的直线相关系数为0.83（N = 40; p < 0.0001）; 2）与其他精液参数的相关性较低; 3）精子结合与生育力之间的关系不是线性的，但阈值图允许识别具有低结合和低生育力的雄性。来自公鸡、火鸡、水牛、人、小鼠、公羊和种马的活动精子以剂量反应方式结合。结合的特点，揭示了扫描和透射电子显微镜。使用这种方法来挑选男性的精液似乎正常的传统模式的分析，但不同的强制性性状的精卵结合可能是有价值的，以避免昂贵的后代测试。
A series of in vitro assays of sperm-egg binding were developed to identify potentially subfertile roosters, Initial assays used either segments of intact hen's egg perivitelline membrane (PVM) placed on a microscope slide or a heat-solubilized extract of PVM (MS-PVM) dried within a flat-bottomed microwell plate, with bound sperm detected with a DNA-specific stain and epifluorescence microscopy. An automated assay was developed using prestained sulfosuccinimidyl-7-amino-4-methylcoumarin-3-acetate-HS-PVM and enumeration of bound sperm with a fluorometric microwell plate reader, Four populations of chickens differing in fertility were evaluated with the following results: 1) the correlation across lines between in vitro sperm binding and fertility was 0.83 (N = 40; p < 0.0001); 2) correlations with other seminal parameters were low; and 3) the relationship between sperm binding and fertility was not linear, bur a threshold plot allowed identification of males with low binding and low fertility Motile sperm from roosters, turkeys, buffs, humans, mice, rams, and stallions bound in a dose-responsive manner. Features of binding were revealed by both scanning and transmission electron microscopy. Use of this assay to cull males whose semen appears normal by traditional modes of analysis but differs in the obligatory trait of sperm-egg binding could be of value to avoid expensive progeny testing.