EBV was detected via in situ hybridization (ISH) using an EBV ISH kit (Leica Microsystems, Wetzlar, Germany). Only cases of EBV‐posDLBCL showing nuclear positivity for EBV‐encoded small RNA (EBER) in most neoplastic cells (>50%) and EBV‐negDLBCL cases without singular EBER‐positive cells were included in the study.
Ebv ish kit
Manufactured by Leica
Sourced in Germany, United States
The EBV ISH kit is a laboratory equipment product designed for the detection and identification of Epstein-Barr virus (EBV) in biological samples using in situ hybridization (ISH) technology. The kit provides the necessary reagents and protocols to perform this specialized analysis.
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4 protocols using ebv ish kit
1
EBV Detection in DLBCL via ISH
2
EBV+ DLBCL Tissue Sample Analysis
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Tissue samples were obtained from 11 patients at the time of their diagnosis with EBV+DLBCL between 2014 and 2017. Table 1 presents the detailed clinical characteristics of the included patients. The 11 tissue samples were evaluated, and then DNA was extracted from formalin-fixed, paraffin-embedded (FFPE) samples.
EBV+ DLBCL was diagnosed according to the 2016 WHO classification of lymphoid neoplasms [1 ]. EBV was detected using an in situ hybridization ISH technique with the EBV ISH kit (Leica Microsystems, Wetzlar, Germany). This study only included cases of EBV+ DLBCL that showed nuclear positivity for the EBV-encoded small RNA (EBER) in a majority (>50%) of neoplastic cells. We excluded EBV+ DLBCL cases in the clinical setting of known immune deficiency.
Informed consent was obtained from the legal guardians of all participating patients before this study, and we had access to information that could identify individual patients during/after data collection. This study was approved by the Ethics Committee of the Institutional Review Board at Sun Yat-sen University Foshan Hospital [File number: L (2014) NO.2].
EBV+ DLBCL was diagnosed according to the 2016 WHO classification of lymphoid neoplasms [1 ]. EBV was detected using an in situ hybridization ISH technique with the EBV ISH kit (Leica Microsystems, Wetzlar, Germany). This study only included cases of EBV+ DLBCL that showed nuclear positivity for the EBV-encoded small RNA (EBER) in a majority (>50%) of neoplastic cells. We excluded EBV+ DLBCL cases in the clinical setting of known immune deficiency.
Informed consent was obtained from the legal guardians of all participating patients before this study, and we had access to information that could identify individual patients during/after data collection. This study was approved by the Ethics Committee of the Institutional Review Board at Sun Yat-sen University Foshan Hospital [File number: L (2014) NO.2].
3
Quantifying Serum CCL3 and EBV Detection
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The serum CCL3 concentration was measured in archived frozen samples of the Samsung Medical Center prospective cohort study (NCT#00822731). Archived serum sample aliquots stored at −80°C were thawed for use in the cytokine assay. The CCL3 concentration was measured in serum using the Procarta cytokine profiling kit (Panomics, San Diego, CA, USA), and all measurements performed in duplicate.
EBER was detected using an EBV ISH kit (Leica Microsystems, Bannockburn, IL, USA). We employed EBV-negative lymphoid tissues and the hybridization mixture without EBV oligonucleotides as the negative control. A positive reaction was defined as >20% cells showing nuclear positivity.
EBER was detected using an EBV ISH kit (Leica Microsystems, Bannockburn, IL, USA). We employed EBV-negative lymphoid tissues and the hybridization mixture without EBV oligonucleotides as the negative control. A positive reaction was defined as >20% cells showing nuclear positivity.
4
Determining Cell of Origin in DLBCL
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The pathology of the DLBCL cases was confirmed by an expert hematopathologist (Y.H.K.) using the WHO classification. To determine the cell of origin of DLBCL, immunohistochemical staining was performed in formalin-fixed paraffin-embedded specimens using a panel of monoclonal antibodies against CD10 (Dakopatts, Copenhagen, Denmark), BCL-6 (Dakopatts), and MUM-1 (Dakopatts). Stained slides were reviewed, and the cell of origin was determined by expert hematopathologists (M.H. and Y.H.K.) according to the results of immunohistochemistry. Thus, patients were classified as having the germinal center B-cell-like (GCB) or non-GCB subtype based on the Hans algorithm, as proposed previously [33 (link)]. EBER was detected using ISH and an EBV ISH kit (Leica Microsystems, Bannockburn, IL, USA). We used EBV-negative lymphoid tissues and the hybridization mixture without EBV oligonucleotides as negative controls. A positive reaction was defined as more than 20% of examined cells showing nuclear positivity, as applied in our previous series [6 (link), 7 (link), 34 (link)].
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