Wagner H J, Wessel M, Jabs W, Smets F, Fischer L, Offner G, Bucsky P
Department of Pediatrics, Medical University of Lübeck, Germany.
Transplantation. 2001 Sep 27;72(6):1012-9. doi: 10.1097/00007890-200109270-00006.
Early diagnosis of Epstein-Barr virus (EBV)-associated posttransplant lymphoproliferative disorder (PTLD) is required to detect a stage of disease that is more likely to respond to treatment. Elevated levels of EBV DNA were found in peripheral blood of patients at the onset of PTLD.
To compare plasma and peripheral blood mononuclear cells (PBMCs) as material for real-time quantitative polymerase chain reaction (RQ-PCR) measurement of Epstein-Barr viral load, we used two sets of primers and probes specific for the BAM HI-K or BAM HI-W region of the EBV genome.
Patients with PTLD had a median viral load of 19,200 EBV genomes/microg DNA (n=9) or 3,225 EBV genomes/100 microl plasma (n=5), being significantly higher compared with immunosuppressed patients with primary (n=9) or reactivated (n=20) EBV infection or immunosuppressed patients without serological signs of active EBV infection (n=67) (P<0.001). Hence, a value of greater than 5,000 EBV genomes/microg PBMC DNA was considered as a diagnostic parameter for PTLD with a sensitivity and specificity of 1.00 or 0.89, respectively. When plasma was analyzed, however, a value of greater than 1,000 EBV genomes/100 microl plasma had both a sensitivity and specificity of 1.00 for the diagnosis of PTLD. During remission of PTLD, viral load was more effectively cleared in plasma compared with PBMCs. In plasma of nonimmunosuppressed individuals, even a qualitative detection of EBV-related sequences was sensitive and specific for the diagnosis of primary EBV infection, whereas for analysis of PBMC DNA a quantitative parameter had to be considered to differentiate healthy individuals (< 100 EBV genomes/microg PBMC DNA) from patients with primary EBV infection (>100 EBV genomes/microg PBMC DNA).
Although both PBMCs and plasma were useful as material for EBV-specific RQ-PCR in immunosuppressed patients and nonimmunosuppressed individuals, the specificity of analysis seemed to be higher if plasma was taken for analysis.
需要对爱泼斯坦-巴尔病毒（EBV）相关的移植后淋巴细胞增生性疾病（PTLD）进行早期诊断，以检测出更可能对治疗产生反应的疾病阶段。在PTLD发病时，患者外周血中EBV DNA水平升高。
为了比较血浆和外周血单个核细胞（PBMC）作为实时定量聚合酶链反应（RQ-PCR）测量EBV病毒载量的材料，我们使用了两组针对EBV基因组BAM HI-K或BAM HI-W区域的引物和探针。
PTLD患者的病毒载量中位数为19,200个EBV基因组/μg DNA（n = 9）或3,225个EBV基因组/100μl血浆（n = 5），与原发性（n = 9）或再激活（n = 20）EBV感染的免疫抑制患者或无活动性EBV感染血清学迹象的免疫抑制患者（n = 67）相比显著更高（P < 0.001）。因此，PBMC DNA中EBV基因组大于5,000个/μg被视为PTLD的诊断参数，其敏感性和特异性分别为1.00或0.89。然而，当分析血浆时，血浆中EBV基因组大于1,000个/100μl对PTLD诊断的敏感性和特异性均为1.00。在PTLD缓解期，与PBMC相比，血浆中的病毒载量清除更有效。在非免疫抑制个体的血浆中，即使是EBV相关序列的定性检测对原发性EBV感染的诊断也具有敏感性和特异性，而对于PBMC DNA分析，必须考虑定量参数以区分健康个体（<100个EBV基因组/μg PBMC DNA）和原发性EBV感染患者（>100个EBV基因组/μg PBMC DNA）。
虽然PBMC和血浆在免疫抑制患者和非免疫抑制个体中都是EBV特异性RQ-PCR的有用材料，但如果采用血浆进行分析，分析的特异性似乎更高。
