AIMS: To develop a rapid, sensitive, and specific non-isotopic in situ hybridisation (NISH) procedure for the detection of Epstein-Barr virus in formalin fixed, paraffin wax embedded tissues. METHODS: Two low molecular weight RNAs, designated EBER-1 and EBER-2 (Epstein-Barr encoded RNA), were used: cells latently infected with EBV secrete large amounts of EBERs. The method uses digoxigenin labelled anti-sense oligonucleotides, corresponding to sequences in EBER-1 and EBER-2. RESULTS: The use of these probes, in conjunction with high temperature microwave denaturation, ensured that the technique was considerably more sensitive than other in situ hybridisation techniques for detecting EBV. Furthermore, the hybridisation signal was morphologically distinct in that only the nucleus and not the nucleolus give a positive signal. No cross-hybridisation was observed with cells infected with other lymphotropic herpes viruses. CONCLUSION: The sensitivity, simplicity, and rapidity of this technique make it ideal for diagnostic use, and for studies investigating the role of this virus in neoplastic disease.
AIMS: To develop a rapid, sensitive, and specific non-isotopic in situ hybridisation (NISH) procedure for the detection of Epstein-Barr virus in formalin fixed, paraffin wax embedded tissues. METHODS: Two low molecular weight RNAs, designated EBER-1 and EBER-2 (Epstein-Barr encoded RNA), were used: cells latently infected with EBV secrete large amounts of EBERs. The method uses digoxigenin labelled anti-sense oligonucleotides, corresponding to sequences in EBER-1 and EBER-2. RESULTS: The use of these probes, in conjunction with high temperature microwave denaturation, ensured that the technique was considerably more sensitive than other in situ hybridisation techniques for detecting EBV. Furthermore, the hybridisation signal was morphologically distinct in that only the nucleus and not the nucleolus give a positive signal. No cross-hybridisation was observed with cells infected with other lymphotropic herpes viruses. CONCLUSION: The sensitivity, simplicity, and rapidity of this technique make it ideal for diagnostic use, and for studies investigating the role of this virus in neoplastic disease.
Authors: Zaher K Otrock; Jad Saab; Georges Aftimos; Fady Nasr; Fadi S Farhat; Saad Khairallah; Gérard Abadjian; Marwan Ghosn; Hassan Sidani; Ahmad Ibrahim; Ayman Tawil; Claude Ghorra; Zarouhie Meguerian; Walid Mokaddem; Walid Dayeh; Ziad Salem; Georges Chahine; Nizar Bitar; Anas Mugharbel; Joseph Makdessi; Christina Khater; Mirna El Hajj; Dany Abi Gerges; Charles Sfeir; Joseph Kattan; Khaled Ibrahim; Michel Saade; Hussein Sadek; Rami A Mahfouz; Mohamed A Kharfan-Dabaja; Ghazi Zaatari; Ali Bazarbachi Journal: Pathol Oncol Res Date: 2013-05-08 Impact factor: 3.201