Wagner H J, Bein G, Bitsch A, Kirchner H
Institute of Immunology and Transfusion Medicine, University of Lübeck Medical School, Germany.
J Clin Microbiol. 1992 Nov;30(11):2826-9. doi: 10.1128/jcm.30.11.2826-2829.1992.
We designed a highly sensitive and specific polymerase chain reaction assay for the detection of Epstein-Barr virus (EBV)-related sequences in B-cell DNA of EBV-seropositive healthy individuals. By using this assay, we were able to amplify at least 10 copies of a plasmid containing the BamHI-W region, which is repeated up to 11 times within the EBV genome, in the presence of 1 microgram of EBV-negative DNA, indicating that one virus genome was detectable from 150,000 cells. In 15 of 16 tested individuals, EBV-related sequences were found frequently when the DNA from 10(6) B lymphocytes was examined and 1 microgram of DNA was used in each polymerase chain reaction. When the results of amplifying the diluted plasmid were used as a semiquantitative standard, the number of EBV genomes detected could be estimated to range between 50 and less than 1 from 10(6) B lymphocytes. The results of our study will provide the basis for further investigations of the characteristics of the latent carrier state in healthy EBV-seropositive individuals, such as the determination of the number of virus copies per cell and expression of antigens.
我们设计了一种高灵敏度和特异性的聚合酶链反应检测方法，用于检测EB病毒血清反应阳性健康个体B细胞DNA中与EB病毒（EBV）相关的序列。通过使用该检测方法，在存在1微克EBV阴性DNA的情况下，我们能够扩增出至少10个含有BamHI-W区域的质粒拷贝，该区域在EBV基因组中重复多达11次，这表明从150,000个细胞中可检测到一个病毒基因组。在16名受试个体中的15名中，当检测来自10⁶个B淋巴细胞的DNA且每个聚合酶链反应使用1微克DNA时，经常发现与EBV相关的序列。当将稀释质粒的扩增结果用作半定量标准时，从10⁶个B淋巴细胞中检测到的EBV基因组数量估计在50至不足1之间。我们的研究结果将为进一步研究健康EBV血清反应阳性个体中潜伏携带状态的特征提供基础，例如确定每个细胞中的病毒拷贝数和抗原表达。
