改良荧光原位杂交三步法在产前诊断绒毛标本中的应用

摘要

目的 对荧光原位杂交方法进行改良,探讨一种更简便高效的杂交方法.方法 采用对照实验收集2012至2013年绍兴市妇幼保健院36份产前诊断绒毛标本,将标本制片后,使用18/X/Y、13/21探针,分别用改良三步法和经典法进行杂交,观察杂交率和荧光信号,并与传统培养结果进行配对t检验.结果 共检测72个杂交区,21/13探针三步法杂交率和杂交信号均明显高于经典法(杂交率99.72％ ±0.42％和85.90％±4.15％,t=20.4,P＜0.01;杂交信号2.58 ±0.50和1.52±0.55,t=7.53,P＜0.01),三步法18/X/Y探针杂效率与信号与经典法差异无统计学意义(杂效率99.57％ ±0.53％和99.70％ ±0.42％,t=1.30,P＞0.05;杂交信号2.22 ±0.42和2.36 ±0.48,t=1.57,P＞0.05);经典法与三步法的荧光原位杂交结果与传统培养结果符合率为100％(36/36).结论 本研究中的改良三步法操作简便,步骤简单、实验效率高、环保,但也存在18/X/Y信号偏散的情况,有待于进一步摸索.%Objective To investigate a simple and efficient fluorescence in situ hybridization method for prenatal diagnosis.Methods Thirty-six cases of chorionic villus samples were hybridized with probes 18/X/Y,13/21 by using traditional cuhure methods and modified three-step methods in controlled experimentation during 2012 to 2013.The hybridization rate and fluorescence signals were analyzed.Results A total of 72 hybrid zone were detected.Probe 21/13 hybridization rate and fluorescence signals of three-step modified methods were higher than that of traditional methods(99.72％ ±0.42％ vs 85.90％ ±4.15％,t =20.4,P ＜0.01; 2.58 ±0.50 vs 1.52 ± 0.55,t =7.53,P ＜0.01).Probe 18/X/Y has the same hybridization rate and fluorescence signals between three-step modified methods and traditional methods (99.57％ ±0.53％ vs99.70％ ±0.42％,t=1.30,P＞0.05; 2.22±0.42 vs2.36±0.48,t=1.57,P＞0.05).The coincidence rate of two methods was 100％ (36/36).Conclusion The modified fluorescence in situ hybridization three-step methods in the study was simple,rapid,effective and environment-friendly.At the same time,it has some defects such as the signals of 18/X/Y was not concentrated enough.Further exploration is needed.