Fluorescent <i>in situ</i> hybridization of EBV and its correlation with qPCR/CCR data in EBV-associated gastric carcinoma.

Fluorescent in situ hybridization (FISH) of EBV-DNA was performed with a specific DNA probe against the full-length EBV genome. Each red signal represents a single copy of the EBV genome. Each nucleus of EBV-infected cell line Namalwa contains one or two signals in each cell (A). Each nucleus of NCC-24 contains a lot of red signals in some cells, while no red signal in others, resulting in the average 3.9/nucleus (B). SNU-719 shows a lot of red signals in each nucleus, 34 on average. There are 7 nuclei in the figure (C). In two EBV-associated gastric carcinoma (EBVaGC) cases, the one shows 9 signals per nucleus (D) and the other 26 signals per nucleus (E). In both figures, several nuclei are aggregated. The graph presents correlation of FISH analysis with qPCR/CCR data (F). The y-axis represents the average number of EBV per nucleus by FISH. The x-axis represents the EBV copy number per nucleus, which was twice the corresponding value of the EBV-CN, assuming no amplification or deletion of GAPDH in the cancer cell nucleus. Dots and bars represent the mean and standard deviation, respectively. For the cases in which 2 × EBV-CN is less than 40, the number of EBV signals in FISH is proportional to 2 × EBV-CN; it gradually comes close to a plateau of 44 when 2 × EBV-CN exceeds 40 (R2 = 0.75 on the Hill curve fitting). A-E: Scale bar is 5 μm. Yellow arrowheads point to a single red signal of the EBV genome in each figure.