论文部分内容阅读
目的建立高效的荧光原位杂交(fluorescence in situ hybridization,FISH)前人精子核去凝集方法和监测手段。方法精子标本经10mmol/L二硫代苏糖醇(dithiothreitol,DTT)处理,使人精子核去凝集,倒置相差显微镜监测精子核膨胀度。标本固定后制片,变性后行FISH分析。结果倒置相差显微镜下,人精子核膨大程度清晰可见,膨大精子细胞边缘清楚并完整。荧光显微镜下,可见精子头部染成蓝色,边缘清楚,背景清晰,精子杂交信号清晰,非特异性杂交少。结论本方法具有简便、高效的特点,精子核解聚程度可控性和可见性强,杂交后荧光信号强、易分辨和非特异性性杂交率低,可用于人精子核FISH前处理。
OBJECTIVE To establish a highly efficient fluorescence in situ hybridization (FISH) decondensation method and monitoring method for human sperm nuclei. Methods Spermatozoa were treated with 10 mmol / L dithiothreitol (DTT) to deaggregate human sperm nuclei, and the degree of sperm nucleus expansion was monitored by inverted phase contrast microscope. Specimens were fixed after production, denatured after FISH analysis. Results inverted phase contrast microscope, the degree of human sperm nuclear enlargement clearly visible, swollen sperm cells clear and complete edge. Under a fluorescence microscope, the head of the sperm is stained blue, the edge is clear, the background is clear, the sperm hybridization signal is clear, and the non-specific hybridization is less. Conclusion The method is simple and efficient. The degree of controllable and visible sperm nucleus depolymerization is strong. After hybridization, the fluorescence signal is strong, easily distinguishable and non-specific hybridization rate is low, which can be used for human sperm nuclear FISH pretreatment.