双色FISH检测人精子染色体非整倍体方法的建立

Establishing the method of two color fluorescence in situ hybridization to detect aneuploidy of human sperm.

目的 建立用双色FISH检测人精子染色体非整倍体的方法。方法 采用双色荧光原位杂交 (FISH)方法取适量精子标本用EDTA/PBS处理 ,然后用二硫苏糖醇 (DTT) ,使精子去凝集。固定后滴片 ,然后与双色荧光直接标记探针杂交。结果 在OLYMPUS荧光显微镜下可以清楚看到精子头部的蓝色杂交信号 ,头部有 1个绿色荧光杂交信号的精子为X染色体精子 (X精子 ) ,有 1个红色荧光杂交信号的精子为Y染色体精子 (Y精子 )。精子头部有 2个荧光杂交信号的精子为染色体数目异常精子。结论 双色荧光原位杂交 (FISH)方法可以用于测定人精子染色体非整倍体率的变化。

Objective:To establish the method of two color fluorescence in situ hybridization(FISH) assay and use it to detect the aneuploidy frequency of human sperm chromosome. Methods: Sperm sample was washed with EDTA/PBS. The sperm heads were decondensed with dithiothreitol (DTT). Then, the sperm nuclei were hybridized with two color direct labeled enumeration probe(chromosome X α- satellite probe (DXZ1) and chromosome Y α- satellite probe (DYZ3). Results:Under the OLYMPUS fluorescence microscope, the hybridization signals in the sperm heads were clearly visible. The sperm with one green hybridization signal was X chromosome sperm (X sperm ), and the sperm with one red hybridization signal in the sperm head was Y chromosome sperm ( Y sperm ). In the case of two hybridization signals in one sperm head, the sperm should be a numerical abnormal one. By using two color FISH with one autosome probe and one sex chromosome probe, the sperm with same color of two hybridization signals in one sperm head could be identified as aneuploidy sperm or diploid sperm. Conclusions: The two color FISH assay may be used to detect the aneuploidy frequency of human sperms.