Assessment of automated DNA extraction coupled with real-time PCR for measuring Epstein–Barr virus load in whole blood, peripheral mononuclear cells and plasma

Evaluation of the Epstein-Barr Virus R-Gene Quantification Kit in Whole Blood with Different Extraction Methods and PCR Platforms

“…11,15 This comparison was made with the EBV QCMD 2006 Proficiency panel and with clinical specimens routinely sent to our institution for EBV DNA quantification. In clinical samples, the correlation for absolute values of EBV DNA load was very good (r ϭ 0.92), and interestingly, this good agreement was also observed in the follow-up of two patients treated with anti-CD20 monoclonal antibodies.…”

“…However, to our knowledge, there is not yet a clear threshold value of EBV DNA load against which a pre-emptive strategy could be warranted to avoid PTLD (ie, reducing immunosuppression and anti-CD20 treatment). 11,16,17 Furthermore, it should be noted here that the value of monitoring EBV DNA load in plasma rather than in whole blood in the transplantation setting is also a subject of controversy. 22,23 In a previous study, 11 we showed high viral loads in all blood compartments (plasma, peripheral blood mononuclear cells, and whole blood) in two patients with proven PTLD, whereas two recent studies 24,25 demonstrated that whole blood was more sensitive for the quantification of EBV DNA in transplant patients than plasma.…”

“…Indeed, this specimen combines all blood compartments that may harbor EBV, and it best reflects the absolute viral burden in the patient's circulation. 11,12 The automation of the nucleic acid purification step before real-time PCR amplification could also simplify and improve the reproducibility of EBV DNA load measurement. 11,13 Because the availability of a variety of in-house PCR assays using different real-time PCR platforms and software may challenge the standardization of EBV DNA load quantification, the development of a versatile commercial PCR amplification assay is another important step in providing clinical laboratories with reliable diagnostic tools.…”

Evaluation of a commercial real-time PCR assay for quantitation of Epstein-Barr virus DNA in different groups of patients

“…Depending on type and stage of disease EBV DNA may also be found in plasma, usually representing fragmented cell-derived material. However, during active infection EBV DNA can be detected for a longer period of time in whole blood than in plasma (Fafi-Kremer et al, 2004).…”

Cellular normalization of viral DNA loads on whole blood improves the clinical management of cytomegalovirus or Epstein Barr virus infections in the setting of pre‐emptive therapy

“…However, it is not possible to differentiate problems of extraction (loss of sample) from partial inhibition of amplification. The association of automated extraction with real-time quantitative PCR has been increasingly developed in recent years for routine diagnosis of CMV and EBV infections [Mengelle et al, 2003a;Fafi-Kremer et al, 2004;Gouarin et al, 2004]. The interest of multiplex, quantitative, and real-time PCR assays that makes it possible to normalize viral load according to the amount of cellular DNA has been underlined previously in a number of studies.…”