Double-step PCR assay to quantify Epstein-Barr viral load in peripheral blood

Molecular Biotechnology
M BergalloR Cavallo

Abstract

Posttransplant lymphoproliferative disorders (PTLD) are a severe complication arising in solid organ transplant patients. A strong correlation between Epstein-Barr virus (EBV) infection, the grade and type of immunosuppression, and the development of PTLD has been recognized. This article describes the development of a double-step polymerase chain reaction (PCR) assay for the quantification of EBV-deoxyribonucleic acid (DNA) to monitor EBV infection. Screening of samples containing >/=10(3) viral genomes/10(5) peripheral blood mononuclear cells (PBMC) or 100 micro L serum by a semiquantitative PCR assay is followed by quantification of the samples containing a high number of viral genomes in a quantitative-competitive (QC)-PCR assay. Screening by semiquantitative PCR selects samples with a high number of viral genomes for use in the more labor-intensive and expensive QC-PCR assay and thus provides a handy means for quantitative DNA analysis of large numbers of samples. Our double-step PCR assay can be employed in EBV viral load measurement in PBMC and serum samples to monitor transplanted patients at risk to develop PTLD.

Citations

Jul 27, 2007·Molecular Biotechnology·Massimiliano BergalloRossana Cavallo
Aug 26, 2006·Digestive and Liver Disease : Official Journal of the Italian Society of Gastroenterology and the Italian Association for the Study of the Liver·A LavagnaA Pera
Jun 6, 2009·Transplant Infectious Disease : an Official Journal of the Transplantation Society·H OmarP Ljungman

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