Xiao Yanping, Chen Meiyue, Sheng Min, Yang Hua, Huang Shuzhen
Shanghai Institute of Medical Genetics, Shanghai Children's Hospital, Shanghai, 200040 P. R. China.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2003 Apr;20(2):147-50.
To establish a high sensitive and specific method of interphase fluorescence in situ hybridization (IFISH) to detect the low-frequency human cells in human/goat xenogeneic models.
Human-specific Y-chromosome satellite DNA CEPY and 17-chromosome satellite DNA p17H8 were used as probes for IFISH. The peripheral blood samples from 2 goats transplanted with human male hematopoietic stem cells (HSC), 1 normal negative goat and 1 normal man were analyzed. The actual FISH efficiency was confirmed by serial dilutions (1/100, 1/500 and 1/1000) of the cell mixture of normal man and normal negative goat. A set of signal scoring criteria was determined to guarantee the stability and reliability of the method.
Positive cell (human cell) frequencies were consistent with the established frequencies for the human/goat cell mixture. The average frequencies of positive cells were 98.60% (CEPY) and 100% (p17H8) for normal man, 0 for normal negative goat, 0.23% (CEPY) and 0.11% (p17H8) for human/goat xenogeneic models. The results demonstrated that low-frequency human cells (male cells confirmed by Y-chromosome probe) existed in human/goat xenogeneic models.
The IFISH developed in this study is of high sensitivity and specificity and can identify the actual frequency of human cells, which offers a direct, sensitive and specific approach to the detection of low-frequency human cells in human/goat xenogeneic models.
建立一种高灵敏度和特异性的间期荧光原位杂交（IFISH）方法，用于检测人/山羊异种模型中的低频人细胞。
使用人特异性Y染色体卫星DNA CEPY和17号染色体卫星DNA p17H8作为IFISH探针。对2只移植了人男性造血干细胞（HSC）的山羊、1只正常阴性山羊和1名正常男性的外周血样本进行分析。通过对正常男性和正常阴性山羊的细胞混合物进行系列稀释（1/100、1/500和1/1000）来确认实际的FISH效率。确定了一套信号评分标准，以保证该方法的稳定性和可靠性。
阳性细胞（人细胞）频率与建立的人/山羊细胞混合物频率一致。正常男性的阳性细胞平均频率为98.60%（CEPY）和100%（p17H8），正常阴性山羊为0，人/山羊异种模型为0.23%（CEPY）和0.11%（p17H8）。结果表明人/山羊异种模型中存在低频人细胞（Y染色体探针确认的男性细胞）。
本研究建立的IFISH具有高灵敏度和特异性，能够鉴定人细胞的实际频率，为检测人/山羊异种模型中的低频人细胞提供了一种直接、灵敏和特异的方法。
