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[摘要] 目的 评价一种新的精子染色方法BASO(改良Diff-Quik)法在精子顶体检测中的应用。方法 收集60例不育症男性精液,用BASO法、改良巴氏法和瑞姬氏混合染色法,同时进行染色,比较其固定和染色时间均数,并分析其精子顶体不完整率检测的差异。结果BASO法固定和染色约需7min,明显短于改良巴氏法和瑞姬氏混合染色法,而精子顶体不完整率检测的差异无显著性。结论 BASO法操作步骤简单省时,可用于临床精子标本的染色,分析精子形态及精子顶体完整性。
[关键词] 染色法;精子顶体;BASO
[中图分类号] R446.5[文献标识码] A [文章编号] 1673-9701(2010)06-29-02
Three Kinds of Staining Methods in Evaluation of Sperm Morphology
WANG ChengZHANG HanwangLIU Qun
Center of Genesiology,Affiliated Tongji Hospital of Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430030,China
[Abstract] Objective To explore the application of a new semen staining method-BASO in the detection of perforatorium. Methods Sixty human semen samples were enrolled into this study. Three semen staining methods were used including BASO method,modified Papanicolaou method and Wright-Giemsa mixed stain. Results The fixation and staining of BASO method needed about 7min,significantly shorter than other two staining methods,and however,there was no significant difference in the detection of the incomplete rate of human perforatorium between these methods. Conclusion BASO method is simple and fast to operate,and can be used for human semen staining,semen morphology analysis and perforatorium integrity.
[Key words] Staining method;Perforatorium;BASO method
精液常规中形态染色和测定顶体酶活性是判断精子生育力很有价值的一种方法,目前改良巴氏染色和瑞-吉氏染色被证明适合用于精子形态染色和顶体着色观察,然而这两种方法操作步骤繁琐,耗时较多,不利于检测报告的及时发放。为此,我们使用了一种新的精子染色方法BASO法,试图通过快捷、有效的精子染色和精子顶体完整率检测程序,为提高人工授精及体外受精环节中精液品质检查效率提供帮助。
1材料与方法
1.1研究对象
60例不育男性均来自本院不育门诊,年龄24～35岁,婚后正常性生活2年以上不育,无外伤及遗传性疾病家族史,无性功能障碍史,体检未发现有明显睾丸、附睾及输精管异常。
1.2标本采集
禁欲3～5d后手淫留取精液在干燥消毒塑料杯内,37℃孵育箱内液化,进行检测。
1.3BASO精子染色法
其含有A、B两种染色缓冲液,配制无特殊要求。具体染色步骤为:涂片空气干燥,然后用等量95%乙醇固定5min,滴1~2滴染液A液覆盖于涂片上染15~20s,用M/15的磷酸盐缓冲液(pH 6.6～6.8)稍冲洗掉染液A液,然后稍甩干缓冲液,滴加2~3滴染液B液覆盖于涂片上染20～30s,流水冲洗,待干燥后涂片即可直接油镜下观察。巴氏染色法和瑞姬氏混合染色法按照说明书步骤操作[1]。每份精子均用三种方法检测,每种方法涂片两张。
1.4精液常规分析
按照WHO精液分析标准进行。正常精子头部呈椭圆形,精子头部长度为4.0～5.0μm,宽为2.5～3.5μm。长宽之比应在1.50～1.75之间。精子长和宽的评估可借助于目镜标尺进行[2]。尾部应是直的、均一的、比中段细、非卷曲的,其长约为45μm,计数200条精子中的正常百分率。
1.5顶体酶结果观察
顶体的界限应清晰,占头部的40%～70%。中段应细,宽度
