[Development and preliminary analysis for EBV clonality using non-RI probe].

Abstract

We develop a method for EBV clonality analysis by southern blot hybridization using Non-RI probes directed to EBV genome. We also tried to use newly developed method for several cells, which have been known as EBV genome carrier, and for a clinically diagnosed specimen from patients of infectious mononucleosis (IM) and chronic active EBV infection(CAEBV). Non-RI probe(directed to terminal repeat of EBV genome, TR probe) were made by following ways; The Xho I fragment(1.9 Kb) of B95-8 cells(carrier of EBV genome) was amplified by PCR, and the product was cloned into pBluescript, and cloned DNA were labeled by digoxigenin(Dig) after purification. The TR probes did not hybridize with DNA from other herpes viruses and Ramos cells(EBV-free cell line). When TR probe used against peripheral blood mononuclear cells(PBMCs) from a CAEBV patient, pattern was monoclonal as same as the case of Raji cells. In the case of B95-8 cells and PBMCs of IM patient, patterns were polyclonal. These results were the same as those of previously reported. This method was possible to detect about 0.3% target Raji's DNA from the mixture of Raji and Ramos cells. Above results indicated that this newly developed method was considered to be superior method in specificity and sensitivity, and also suggested that it was useful for analyzing the clonality of EBV infected cells in daily clinical examinations without radiological equipment and facilities.